83 results on '"Shuangjun Lin"'
Search Results
2. Oxidative Indole Dearomatization for Asymmetric Furoindoline Synthesis by a Flavin‐Dependent Monooxygenase Involved in the Biosynthesis of Bicyclic Thiopeptide Thiostrepton
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Wen Liu, Xiao-Wei Liang, Jian Wang, Shu-Li You, Jiang Tao, Shuangjun Lin, Yufeng Xue, and Zhi Lin
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Indole test ,Indoles ,Molecular Structure ,Bicyclic molecule ,010405 organic chemistry ,Chemistry ,Stereochemistry ,Enantioselective synthesis ,Total synthesis ,General Chemistry ,Flavin group ,General Medicine ,010402 general chemistry ,Thiostrepton ,01 natural sciences ,Catalysis ,Mixed Function Oxygenases ,0104 chemical sciences ,Stereocenter ,chemistry.chemical_compound ,Flavins ,Stereoselectivity ,Oxidation-Reduction - Abstract
The interest in indole dearomatization, which serves as a useful tool in the total synthesis of related alkaloid natural products, has recently been renewed with the intention of developing new methods efficient in both yield and stereoselective control. Here, we report an enzymatic approach for the oxidative dearomatization of indoles in the asymmetric synthesis of a variety of furoindolines with a vicinal quaternary carbon stereogenic center. This approach depends on the activity of a flavin-dependent monooxygenase, TsrE, which is involved in the biosynthesis of bicyclic thiopeptide antibiotic thiostrepton. TsrE catalyzes 2,3-epoxidation and subsequent epoxide opening in a highly enantioselective manner during the conversion of 2-methyl-indole-3-acetic acid or 2-methyl-tryptophol to furoindoline , with up to > 99% conversion and > 99% ee under mild reaction conditions. Complementing current chemical methods for oxidative indole dearomatization, the TsrE activity-based approach enriches the toolbox in the asymmetric synthesis of products possessing a furoindoline skeleton.
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- 2021
3. Characterization of Pyridomycin B Reveals the Formation of Functional Groups in Antimycobacterial Pyridomycin
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Zixing Deng, Shuangjun Lin, Tingting Huang, Maolong Wei, Lin Chen, Zihua Zhou, and Zhihong Xiao
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Alanine ,chemistry.chemical_classification ,Ecology ,biology ,Chemistry ,medicine.drug_class ,Stereochemistry ,INHA ,Active site ,Mycobacterium tuberculosis ,Antimycobacterial ,Applied Microbiology and Biotechnology ,Streptomyces ,Nonribosomal peptide ,Polyketide synthase ,Catalytic triad ,biology.protein ,medicine ,Moiety ,Peptide Synthases ,Oxidoreductases ,Oligopeptides ,Polyketide Synthases ,Food Science ,Biotechnology - Abstract
Pyridomycin, a cyclodepsipeptide with potent antimycobacterial activity, specifically inhibits the InhA enoyl reductase of Mycobacterium tuberculosis. Structure-activity relationship studies indicated that the enolic acid moiety in the pyridomycin core system is an important pharmacophoric group, and the natural configuration of the C-10 hydroxyl contributes to the bioactivity of pyridomycin. The ring structure of pyridomycin was generated by the nonribosomal peptide synthetase (NRPS) and polyketide synthase (PKS) hybrid system (PyrE-PyrF-PyrG). Bioinformatics analysis reveals that short-chain dehydrogenase/reductase (SDR) family protein Pyr2 functions as a 3-oxoacyl acyl carrier protein (ACP) reductase in the pyridomycin pathway. Inactivation of pyr2 resulted in accumulation of pyridomycin B, a new pyridomycin analogue featured with enol moiety in pyridyl alanine moiety and a saturated 3-methylvaleric acid group. The elucidated structure of pyridomycin B suggests that rather than functioning as a post-tailoring enzyme, Pyr2 catalyzes ketoreduction to form the C-10 hydroxyl group in pyridyl alanine moiety and the double bond formation of the enolic acid moiety derived from isoleucine when the intermediate assembled by PKS-NRPS machinery is still tethered to the last NRPS module in a special energy-saving manner. Ser-His-Lys residues constitute the active site of Pyr2, which is different from the typically conserved Tyr-based catalytic triad in the majority of SDRs. Site-directed mutation identified that His154 in the active site is a critical residue for pyridomycin B production. These findings will improve our understanding of pyridomycin biosynthetic logic, identify the missing link for the double bound formation of enol ester in pyridomycin, and enable the creation of chemical diversity of pyridomycin derivatives. IMPORTANCE Tuberculosis (TB) is one of the world’s leading causes of death by infection. Recently, pyridomycin, the antituberculous natural product from Streptomyces has garnered considerable attention for being determined as a target inhibitor of InhA enoyl reductase of Mycobacterium tuberculosis. In this study, we report a new pyridomycin analogue from mutant HTT12, demonstrate the essential role of a previously ignored gene pyr2 in pyridomycin biosynthetic pathway, and imply that Pyr2 functions as a trans ketoreductase (KR) contributing to the formation of functional groups of pyridomycin utilizing a distinct catalytic mechanism. As enol moiety are important for pharmaceutical activities of pyridomycin, our work would expand our understanding of the mechanism of SDR family proteins and set the stage for future bioengineering of new pyridomycin derivatives.
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- 2022
4. Functional Genome Mining Reveals a Class V Lanthipeptide Containing a<scp>d</scp>‐Amino Acid Introduced by an F420H2‐Dependent Reductase
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Zhuo Cheng, Fei Zhang, Ghader Bashiri, Jing Wang, Jiali Hong, Sheng-Xiong Huang, Zixin Deng, Yemin Wang, Shuangjun Lin, Min Xu, Lijun Xu, Meifeng Tao, and Xuefei Chen
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chemistry.chemical_classification ,010405 organic chemistry ,Stereochemistry ,General Chemistry ,General Medicine ,Lantibiotics ,Reductase ,010402 general chemistry ,01 natural sciences ,Genome ,Catalysis ,0104 chemical sciences ,chemistry.chemical_compound ,Enzyme ,chemistry ,Biosynthesis ,Dehydroalanine ,Gene cluster ,Lanthionine - Abstract
Lantibiotics are a type of ribosomally synthesized and post-translationally modified peptides (termed lanthipeptides) with often potent antimicrobial activity. Herein, we report the discovery of a new lantibiotic, lexapeptide, using the library expression analysis system (LEXAS) approach. Lexapeptide has rare structural modifications, including N-terminal (N,N)-dimethyl phenylalanine, C-terminal (2-aminovinyl)-3-methyl-cysteine, and d-Ala. The characteristic lanthionine moiety in lexapeptide is formed by three proteins (LxmK, LxmX, and LxmY), which are distinct from enzymes known to be involved in lanthipeptide biosynthesis. Furthermore, a novel F420 H2 -dependent reductase (LxmJ) from the lexapeptide biosynthetic gene cluster (BGC) is identified to catalyze the reduction of dehydroalanine to install d-Ala. Our findings suggest that lexapeptide is the founding member of a new class of lanthipeptides that we designate as class V. We also identified further class V lanthipeptide BGCs in actinomycetes and cyanobacteria genomes, implying that other class V lantibiotics await discovery.
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- 2020
5. Bioconversion of a ganoderic acid 3-hydroxy-lanosta-8,24-dien-26-oic acid by a crude enzyme from Ganoderma lucidum
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Jian-Jiang Zhong, Han Xiao, Shuangjun Lin, Siqin Cai, and Xiaozheng Wang
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chemistry.chemical_classification ,Chromatography ,Bioconversion ,Lanosterol ,Ganoderic acid ,Substrate (chemistry) ,Bioengineering ,Dehydrogenase ,Applied Microbiology and Biotechnology ,Biochemistry ,chemistry.chemical_compound ,Enzyme ,chemistry ,Biocatalysis ,Ganoderma lucidum - Abstract
Ganoderic acids (GAs) are oxygenated lanostane-type triterpenoids from the traditional medicinal mushroom Ganoderma lucidum and of significant biological activities. Although a ganoderic acid 3-hydroxy-lanosta-8,24-dien-26-oic acid (HLDOA) was found to be biosynthesized from lanosterol, further post-modification of HLDOA is yet unclear. In this work, by using HLDOA as the substrate and a crude enzyme from G. lucidum as the biocatalyst, we observed a new peak in liquid chromatography from the reaction system. The product was purified and identified to be 3-oxo-lanosta-8,24-dien-26-oic acid (OLDOA), which may be converted from HLDOA by a putative dehydrogenase of G. lucidum. The work is useful to future manufacture of GAs as well as their biosynthetic pathway elucidation.
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- 2020
6. Antimicrobial Activity with Enhanced Mechanical Properties in Phenylalanine-Based Chiral Coassembled Hydrogels: The Influence of Pyridine Hydrazide Derivatives
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Chuanliang Feng, Chao Xing, Vincent Mukwaya, Li Yang, Shuangjun Lin, Auphedeous Yinme Dang-i, Tingting Huang, Nabila Mehwish, and Xiaoqiu Dou
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Biochemistry (medical) ,Biomedical Engineering ,Supramolecular chemistry ,Phenylalanine ,General Chemistry ,Hydrazide ,Antimicrobial ,Combinatorial chemistry ,Biomaterials ,Bipyridine ,chemistry.chemical_compound ,chemistry ,Pyridine ,Self-healing hydrogels ,Chirality (chemistry) - Abstract
Hydrazide derivatives are known to display a wide range of biological properties including antimicrobial activities, hence making them desirable candidates for soft biomaterials. Herein, we report chiral supramolecular coassembled hydrogels obtained from two phenylalanine gelators (L/DPF and B2L/D) and two dicarbohydrazide molecules (pyridine-2,6-dicarbohydrazide (PDH) and (2,2'-bipyridine)-5,5'-dicarbohydrazide (BDH)) that exhibited enhanced mechanical properties, chirality modulation, and antimicrobial activity. Four lines of coassembled hydrogels were obtained (
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- 2020
7. Biosynthetic access to the rare antiarose sugar via an unusual reductase-epimerase
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Jing Yang, Sheng-Xiong Huang, Meifeng Tao, Geoff P. Horsman, Li Wang, Yijun Yan, Xiaowei Guo, Shuangjun Lin, Dongdong Xu, and Zhiyin Yu
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chemistry.chemical_classification ,010405 organic chemistry ,Biological activity ,General Chemistry ,Reductase ,010402 general chemistry ,01 natural sciences ,Tropolone ,0104 chemical sciences ,chemistry.chemical_compound ,Enzyme ,Aglycone ,chemistry ,Biosynthesis ,Biochemistry ,Epimer ,Gene - Abstract
Rubrolones, isatropolones, and rubterolones are recently isolated glycosylated tropolonids with notable biological activity. They share similar aglycone skeletons but differ in their sugar moieties, and rubterolones in particular have a rare deoxysugar antiarose of unknown biosynthetic provenance. During our previously reported biosynthetic elucidation of the tropolone ring and pyridine moiety, gene inactivation experiments revealed that RubS3 is involved in sugar moiety biosynthesis. Here we report the in vitro characterization of RubS3 as a bifunctional reductase/epimerase catalyzing the formation of TDP-d-antiarose by epimerization at C3 and reduction at C4 of the key intermediate TDP-4-keto-6-deoxy-d-glucose. These new findings not only explain the biosynthetic pathway of deoxysugars in rubrolone-like natural products, but also introduce RubS3 as a new family of reductase/epimerase enzymes with potential to supply the rare antiarose unit for expanding the chemical space of glycosylated natural products.
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- 2020
8. A Novel Mice Model of Catecholaminergic Polymorphic Ventricular Tachycardia Generated by CRISPR/Cas9
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Changliang Hou, Xue Jiang, Q. Qiu, Jiancheng Zheng, Sun Chen, Yunting Zhang, M. Xu, Shuangjun Lin, Lijian Xie, and x. tingting
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medicine.medical_specialty ,business.industry ,chemistry.chemical_element ,Calcium ,medicine.disease ,Ventricular tachycardia ,Catecholaminergic polymorphic ventricular tachycardia ,Ryanodine receptor 2 ,Calcium in biology ,Sudden cardiac death ,Contractility ,Endocrinology ,chemistry ,Internal medicine ,Ca2+/calmodulin-dependent protein kinase ,cardiovascular system ,medicine ,business - Abstract
Catecholaminergic polymorphic ventricular tachycardia (CPVT) has been considered as one of the most important causes of children’s sudden cardiac death. Mutations in the genes for RyR2 and CASQ2, two mainly subtypes of CPVT, have been identified. However, the mutation in the gene of TECRL was rarely reported, which could be another genetic cause of CPVT. We evaluated myocardial contractility, electrophysiology, calcium handling in Tecrl knockout (Tecrl KO) mice and human induced pluripotent stem cell-derived cardiomyocytes. Immediately after epinephrine plus caffeine injection, Tecrl KO mice showed much more multiple premature ventricular beats and ventricular tachycardia. The Tecrl KO mice demonstrate CPVT phenotypes. Mechanistically, intracellular calcium amplitude was reduced, while time to baseline of 50 was increased in acute isolated cardiomyocytes. RyR2 protein levels decreased significantly upon cycloheximide treatment in TECRL deficiency cardiomyocytes. Overexpression of TECRL and KN93 can partially reverse cardiomyocytes calcium dysfunction, and this is p-CaMKII/CaMKII dependent. Therefore, a new CPVT mouse model was constructed. We propose a previously unrecognized mechanism of TECRL and provide support for the therapeutic targeting of TECRL in treating CPVT.
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- 2021
9. Biosynthetic access to the rare antiarose sugar
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Yijun, Yan, Jing, Yang, Li, Wang, Dongdong, Xu, Zhiyin, Yu, Xiaowei, Guo, Geoff P, Horsman, Shuangjun, Lin, Meifeng, Tao, and Sheng-Xiong, Huang
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Chemistry - Abstract
Rubrolones, isatropolones, and rubterolones are recently isolated glycosylated tropolonids with notable biological activity. They share similar aglycone skeletons but differ in their sugar moieties, and rubterolones in particular have a rare deoxysugar antiarose of unknown biosynthetic provenance. During our previously reported biosynthetic elucidation of the tropolone ring and pyridine moiety, gene inactivation experiments revealed that RubS3 is involved in sugar moiety biosynthesis. Here we report the in vitro characterization of RubS3 as a bifunctional reductase/epimerase catalyzing the formation of TDP-d-antiarose by epimerization at C3 and reduction at C4 of the key intermediate TDP-4-keto-6-deoxy-d-glucose. These new findings not only explain the biosynthetic pathway of deoxysugars in rubrolone-like natural products, but also introduce RubS3 as a new family of reductase/epimerase enzymes with potential to supply the rare antiarose unit for expanding the chemical space of glycosylated natural products., Rubrolones, isarubrolones, and rubterolones are recently isolated glycosylated tropolonids with notable biological activity.
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- 2021
10. Characterization of Lysozyme-Like Effector TseP Reveals the Dependence of Type VI Secretion System (T6SS) Secretion on Effectors in Aeromonas dhakensis Strain SSU
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Tong-Tong Pei, Tao G. Dong, Weiliang Xiong, Shuangjun Lin, Zeng-Hang Wang, Li-Li Wu, Ping Xu, and Xiaoye Liang
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Mutant ,Cell ,Virulence ,medicine.disease_cause ,Applied Microbiology and Biotechnology ,Dictyostelium discoideum ,03 medical and health sciences ,Bacterial Proteins ,Phagocytosis ,Cell Wall ,medicine ,Escherichia coli ,Environmental Microbiology ,Secretion ,Dictyostelium ,030304 developmental biology ,Type VI secretion system ,0303 health sciences ,Mutation ,Ecology ,biology ,030306 microbiology ,Chemistry ,Effector ,Type VI Secretion Systems ,biology.organism_classification ,Cell biology ,medicine.anatomical_structure ,Muramidase ,Aeromonas ,Food Science ,Biotechnology - Abstract
The type VI secretion system (T6SS) is a widespread weapon employed by Gram-negative bacteria for interspecies interaction in complex communities. Analogous to a contractile phage tail, the double-tubular T6SS injects toxic effectors into prokaryotic and eukaryotic neighboring cells. Although effectors dictate T6SS functions, their identities remain elusive in many pathogens. Here, we report the lysozyme-like effector TseP in Aeromonas dhakensis, a waterborne pathogen that can cause severe gastroenteritis and systemic infection. Using secretion, competition, and enzymatic assays, we demonstrate that TseP is a T6SS-dependent effector with cell wall-lysing activities, and TsiP is its cognate immunity protein. Triple deletion of tseP and two known effector genes, tseI and tseC, abolished T6SS-mediated secretion, while complementation with any single effector gene partially restored bacterial killing and Hcp secretion. In contrast to whole-gene deletions, the triple-effector inactivation in the 3eff(c) mutant abolished antibacterial killing but not T6SS secretion. We further demonstrate that the 3eff(c) mutation abolished T6SS-mediated toxicity of SSU to Dictyostelium discoideum amoebae, suggesting that the T6SS physical puncture is nontoxic to eukaryotic cells. These data highlight not only the necessity of possessing functionally diverse effectors for survival in multispecies communities but also that effector inactivation would be an efficient strategy to detoxify the T6SS while preserving its delivery efficiency, converting the T6SS to a platform for protein delivery to a variety of recipient cells. IMPORTANCE Delivery of cargo proteins via protein secretion systems has been shown to be a promising tool in various applications. However, secretion systems are often used by pathogens to cause disease. Thus, strategies are needed to detoxify secretion systems while preserving their efficiency. The T6SS can translocate proteins through physical puncture of target cells without specific surface receptors and can target a broad range of recipients. In this study, we identified a cell wall-lysing effector, and by inactivating it and the other two known effectors, we have built a detoxified T6SS-active strain that may be used for protein delivery to prokaryotic and eukaryotic recipient cells.
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- 2021
11. Enzymatic Pyran Formation Involved in Xiamenmycin Biosynthesis
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Xu-Liang Bu, Jun Xu, Jianting Zheng, Min-Juan Xu, Ting Zhou, Jing-Yi Weng, Shuangjun Lin, Bei-Bei He, and Yi-Lei Zhao
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chemistry.chemical_compound ,chemistry ,Biosynthesis ,Pyran ,Stereochemistry ,Furan ,Epoxide ,General Chemistry ,Monooxygenase ,Ring (chemistry) ,Cyclase ,Catalysis ,Benzopyran - Abstract
The pyran ring is a very common structural unit of many natural, bioactive molecules that are widely found in plants, bacteria, and fungi. However, the enzymatic processes by which many of these pyran-containing molecules are formed are unclear. Herein, we report the construction of the pyran ring catalyzed by the cooperation of a flavin-dependent monooxygenase, XimD, and a SnoaL-like cyclase, XimE, in the biosynthesis of xiamenmycins. XimD catalyzes the formation of an epoxide intermediate that spontaneously transforms to furan and pyran products (43:1) in vitro. XimE then catalyzes the formation of the pyran ring in a 6-endo configuration from the epoxide to yield a benzopyran, xiamenmycin B. Further, we obtained the crystallographic structure of XimE, with and without product, which suggests a synergistic mechanism in which a group of four residues (Y46–Y90–H102–E136) acts cooperatively as the general acid and base. Subsequent structure-based analysis of possible viable substrates indicates that both X...
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- 2019
12. Biosynthesis of squalene-type triterpenoids in Saccharomyces cerevisiae by expression of CYP505D13 from Ganoderma lucidum
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Xin Song, Han Xiao, Shangwen Luo, Wenfang Wang, Xiaozheng Wang, and Shuangjun Lin
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0106 biological sciences ,0301 basic medicine ,lcsh:Biotechnology ,Saccharomyces cerevisiae ,Biomedical Engineering ,Heterologous ,Ganoderma lucidum ,lcsh:Chemical technology ,01 natural sciences ,lcsh:Technology ,03 medical and health sciences ,chemistry.chemical_compound ,Squalene ,Biosynthesis ,010608 biotechnology ,Cytochrome P450s (CYPs) ,lcsh:TP248.13-248.65 ,lcsh:TP1-1185 ,Synthetic biology ,chemistry.chemical_classification ,biology ,Renewable Energy, Sustainability and the Environment ,lcsh:T ,Squalene-type triterpenoids (STs) ,biology.organism_classification ,Bioproduction ,In vitro ,030104 developmental biology ,Enzyme ,chemistry ,Biochemistry ,Fermentation ,Food Science ,Biotechnology - Abstract
Background Squalene-type triterpenoids (STs) are a class of linearized triterpenoids with significant bioactivities, including anti-cancer, anti-oxidative, and anti-inflammatory activities. The efficient biosynthesis of STs has gained increasing attention. Results Using Saccharomyces cerevisiae as a heterologous host, we discovered that overexpression of CYP505D13 from Ganoderma lucidum, a famous medicinal mushroom capable of producing various triterpenoids as secondary metabolites, enables the engineered S. cerevisiae strain to produce two new STs, 4,8-dihydroxy-22,23-oxidosqualene (ST-1), 8-hydroxy-2,3;22,23-squalene dioxide (ST-2), and a known ST, 2,3; 22,23-squalene dioxide (ST-3), at the respective titers of 3.28 mg/L, 13.77 mg/L, and 12.23 mg/L after 59 h fermentation. Furthermore, our in vitro enzymatic assay implies that CYP505D13 is involved in the formation of ST-3. Conclusions This study provides a promising alternative to discover STs and facilitate their efficient bioproduction.
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- 2019
13. The molecular basis for the intramolecular migration (NIH shift) of the carboxyl group duringpara-hydroxybenzoate catabolism
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Jim C. Spain, Ying Xu, Huan Zhao, Ning-Yi Zhou, and Shuangjun Lin
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0301 basic medicine ,Stereochemistry ,Gentisates ,030106 microbiology ,Substituent ,Parabens ,Biology ,Hydroxylation ,Thioester ,Microbiology ,03 medical and health sciences ,chemistry.chemical_compound ,Thioesterase ,Molecular Biology ,Biotransformation ,chemistry.chemical_classification ,DNA ligase ,Brevibacillus ,Acetyl-CoA ,Enzymes ,030104 developmental biology ,chemistry ,Hydroxybenzoate ,NIH shift ,lipids (amino acids, peptides, and proteins) ,Metabolic Networks and Pathways - Abstract
The NIH shift is a chemical rearrangement in which a substituent on an aromatic ring undergoes an intramolecular migration, primarily during an enzymatic hydroxylation reaction. The molecular mechanism for the NIH shift of a carboxyl group has remained a mystery for 40 years. Here, we elucidate the molecular mechanism of the reaction in the conversion of para-hydroxybenzoate (PHB) to gentisate (GA, 2, 5-dihydroxybenzoate). Three genes (phgABC) from the PHB utilizer Brevibacillus laterosporus PHB-7a encode enzymes (p-hydroxybenzoyl-CoA ligase, p-hydroxybenzoyl-CoA hydroxylase and gentisyl-CoA thioesterase, respectively) catalyzing the conversion of PHB to GA via a route involving CoA thioester formation, hydroxylation concomitant with a 1, 2-shift of the acetyl CoA moiety and thioester hydrolysis. The shift of the carboxyl group was established rigorously by stable isotopic experiments with heterologously expressed phgABC, converting 2, 3, 5, 6-tetradeutero-PHB and [carboxyl-13 C]-PHB to 3, 4, 6-trideutero-GA and [carboxyl-13 C]-GA respectively. This is distinct from the NIH shifts of hydrogen and aceto substituents, where a single oxygenase catalyzes the reaction without the involvement of a thioester. The discovery of this three-step strategy for carboxyl group migration reveals a novel role of the CoA thioester in biochemistry and also illustrates the diversity and complexity of microbial catabolism in the carbon cycle.
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- 2018
14. Metabolism analysis of 17α-ethynylestradiol by Pseudomonas citronellolis SJTE-3 and identification of the functional genes
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Yali Fu, Yanqiu Wang, Wanli Peng, Zixin Deng, Xin Sun, Rubing Liang, Shuangjun Lin, and Ben Jia
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Environmental Engineering ,Ethanol ,biology ,Health, Toxicology and Mutagenesis ,Cytochrome P450 ,Estrogens ,Dehydrogenase ,Metabolism ,Ethinyl Estradiol ,biology.organism_classification ,Pollution ,chemistry.chemical_compound ,Biodegradation, Environmental ,chemistry ,Biochemistry ,Pseudomonas ,biology.protein ,Environmental Chemistry ,Yeast extract ,Pseudomonas citronellolis ,Energy source ,Waste Management and Disposal ,Gene - Abstract
Synthetic estrogens are the most hazardous and persistent environmental estrogenic contaminants, with few reports on their biodegradation. Pseudomonas citronellolis SJTE-3 degraded natural steroids efficiently and metabolized 17α-ethynylestradiol (EE2) with the addition of different easily used energy sources (glucose, peptone, ethanol, yeast extract, fulvic acid and ammonia). Over 92% of EE2 (1 mg/L) and 55% of EE2 (10 mg/L) in culture were removed in seven days with the addition of 0.1% ethanol, and the EE2-biotransforming efficiency increased with the increasing ethanol concentrations. Two novel intermediate metabolites of EE2 (C22H22O and C18H34O2) were identified with high-performance liquid chromatography (HPLC) and GC-Orbitrap/MS. Comparative analysis and genome mining revealed strain SJTE-3 contained a unique genetic basis for EE2 metabolism, and the putative EE2-degrading genes exhibited dispersed distribution. The EE2 metabolism of strain SJTE-3 was inducible and the transcription of eight genes were significantly induced by EE2. Three genes (sdr3, yjcH and cyp2) encoding a short-chain dehydrogenase, a membrane transporter and a cytochrome P450 hydroxylase, respectively, were vital for EE2 metabolism in strain SJTE-3; their over-expression accelerated EE2 metabolic processes and advanced the generation of intermediate metabolites. This work could promote the study of bacterial EE2 metabolism mechanisms and facilitate efficient bioremediation for EE2 pollution.
- Published
- 2022
15. Spot 42 RNA regulates putrescine catabolism in Escherichia coli by controlling the expression of puuE at the post-transcription level
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Wanli Peng, Guochen Wan, Zixin Deng, Xin Sun, Shuangjun Lin, Ruyan Li, and Rubing Liang
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Spot 42 RNA ,0303 health sciences ,Binding Sites ,Transcription, Genetic ,030306 microbiology ,Chemistry ,Catabolism ,Catabolite repression ,Repressor ,General Medicine ,Gene Expression Regulation, Bacterial ,Applied Microbiology and Biotechnology ,Microbiology ,03 medical and health sciences ,chemistry.chemical_compound ,RNA, Bacterial ,Biochemistry ,Transcription (biology) ,Putrescine ,Escherichia coli ,Animals ,Energy source ,Gene ,030304 developmental biology - Abstract
Putrescine, a typical polyamine compound important for cell growth and stress resistance, can be utilized as an energy source. However, the regulation of its catabolism is unclear. Here the small RNA (sRNA) Spot 42, an essential regulator of carbon catabolite repression (CCR), was confirmed to participate in the post-transcriptional regulation of putrescine catabolism in Escherichia coli. Its encoding gene spf exclusively exists in the γ-proteobacteria and contains specific binding sites to the 5'-untranslated regions of the puuE gene, which encodes transaminase in the glutamylated putrescine pathway of putrescine catabolism converting γ-aminobutyrate (GABA) into succinate semialdehyde (SSA). The transcription of the spf gene was induced by glucose, inhibited by putrescine, and unaffected by PuuR, the repressor of puu genes. Excess Spot 42 repressed the expression of PuuE significantly in an antisense mechanism through the direct and specific base-pairing between the 51`-57 nt of Spot 42 and the 5'-UTR of puuE. Interestingly, Spot 42 mainly influenced the stability of the puuCBE transcript. This work revealed the regulatory role of Spot 42 in putrescine catabolism, in the switch between favorable and non-favorable carbon source utilization, and in the balance of metabolism of carbon and nitrogen sources.
- Published
- 2020
16. Naphthoquinone-Based Meroterpenoids from Marine-Derived Streptomyces sp. B9173
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Xiaozheng Wang, Shuangjun Lin, Tingting Huang, Xinqian Shen, and Zixin Deng
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Aquatic Organisms ,Stereochemistry ,natural products ,Cell Survival ,lcsh:QR1-502 ,naphthoquinone ,Antineoplastic Agents ,Microbial Sensitivity Tests ,010402 general chemistry ,01 natural sciences ,Biochemistry ,Streptomyces ,lcsh:Microbiology ,Article ,chemistry.chemical_compound ,Residue (chemistry) ,Biosynthesis ,Prenylation ,Humans ,meroterpenoid ,Molecular Biology ,Cell Proliferation ,Biological Products ,biology ,Molecular Structure ,010405 organic chemistry ,Terpenes ,biology.organism_classification ,Naphthoquinone ,0104 chemical sciences ,Anti-Bacterial Agents ,Biosynthetic Pathways ,chemistry ,A549 Cells ,Antibacterial activity ,Two-dimensional nuclear magnetic resonance spectroscopy ,flaviogeranin family ,Bacteria ,HeLa Cells ,Naphthoquinones - Abstract
Naphthoquinone-based meroterpenoids are hybrid polyketide-terpenoid natural products with chemical diversity and a broad range of biological activities. Here, we report the isolation of a group of naphthoquinone-containing compounds from Streptomyces sp. B9173, and their structures were elucidated by using a combination of spectroscopic techniques, including 1D, 2D NMR, and high-resolution mass (HRMS) analysis. Seven flaviogeranin congeners or intermediates, three of which were new, have been derived from common naphthoquinone backbone and subsequent oxidation, methylation, prenylation, and amino group incorporation. Both flaviogeranin B1 (1) and B (2) contain an amino group which was incorporated into the C8 of 1,3,6,8-terhydroxynaphthalene (THN). Flaviogeranin D (3) contains an intact C-geranylgeranyl residue attached to the C2 of THN, while the O-geranylgeranyl group of 2 links with the hydroxyl on the C2 site of THN. Four compounds were selected and tested for antibacterial activity and cytotoxicity, with 3 and flaviogeranin C2 (5) displaying potent activity against selected bacteria and cancer cell lines. In light of the structure features of isolated compounds and the biosynthetic genes, a biosynthetic pathway of naphthoquinone-based flaviogeranins has been proposed. These isolated compounds not only extend the structural diversity but also represent new insights into the biosynthesis of naphthoquinone-based meroterpenoids.
- Published
- 2020
17. Characterization of the Phenanthrene-Degrading Sphingobium yanoikuyae SJTF8 in Heavy Metal Co-Existing Liquid Medium and Analysis of Its Metabolic Pathway
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Chong Yin, Wanli Peng, Hua Qiu, Zixin Deng, Shuangjun Lin, Weiliang Xiong, and Rubing Liang
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0301 basic medicine ,Microbiology (medical) ,inorganic chemicals ,phenanthrene ,polycyclic aromatic hydrocarbons ,010501 environmental sciences ,01 natural sciences ,Microbiology ,Sphingobium ,Article ,03 medical and health sciences ,chemistry.chemical_compound ,Bioremediation ,bioremediation ,Virology ,lcsh:QH301-705.5 ,0105 earth and related environmental sciences ,Naphthalene ,Anthracene ,tolerance ,biology ,Strain (chemistry) ,Chemistry ,Phenanthrene ,heavy metal ,biology.organism_classification ,Metabolic pathway ,030104 developmental biology ,Sphingobium yanoikuyae SJTF8 ,lcsh:Biology (General) ,Dibenzothiophene ,Environmental chemistry - Abstract
Polycyclic aromatic hydrocarbons (PAHs) are common organic pollutants with great carcinogenic threaten, and metal/PAH-contaminated environments represent one of the most difficult remedial challenges. In this work, Sphingobium yanoikuyae SJTF8 was isolated and identified with great and stable PAH-degrading efficiency even under stress conditions. It could utilize typical PAHs (naphthalene, phenanthrene, and anthracene) and heterocyclic and halogenated aromatic compounds (dibenzothiophene and 9-bromophenanthrene) as the sole carbon source. It could degrade over 98% of 500 mg/L phenanthrene in 4 days, and the cis-3,4-dihydrophenanthrene-3,4-diol was the first-step intermediate. Notably, strain SJTF8 showed great tolerance to heavy metals and acidic pH. Supplements of 0.30 mM of Cu2+, 1.15 mM of Zn2+, and 0.01 mM of Cd2+ had little effect on its cell growth and phenanthrene degradation, phenanthrene of 250 mg/L could still be degraded completely in 48 h. Further, the whole genome sequence of S. yanoikuyae SJTF8 was obtained, and three plasmids were found. The potential genes participating in stress-tolerance and PAH-degradation were annotated and were found mostly distributed in plasmids 1 and 2. Elimination of plasmid 2 resulted in the loss of the PAH-degradation ability. On the basis of genome mining results, the possible degrading pathway and the metabolites of S. yanoikuyae SJTF8 to phenanthrene were predicted.
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- 2020
- Full Text
- View/download PDF
18. A novel streptonigrin type alkaloid from the Streptomyces flocculus CGMCC 4.1223 mutant ΔstnA/Q2
- Author
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Xiaozheng Wang, Zixin Deng, Shuangjun Lin, Tingting Huang, and Fei Xu
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Methyltransferase ,Double mutant ,010405 organic chemistry ,Chemistry ,Stereochemistry ,Alkaloid ,Organic Chemistry ,Mutant ,Plant Science ,01 natural sciences ,Biochemistry ,0104 chemical sciences ,Analytical Chemistry ,nervous system diseases ,010404 medicinal & biomolecular chemistry ,chemistry.chemical_compound ,Streptonigrin ,surgical procedures, operative ,nervous system ,Streptomyces flocculus ,Gene cluster - Abstract
Streptonigrin (STN) is a highly functionalized aminoquinone alkaloid with broad and potent antitumor activities. Previously, the biosynthetic gene cluster of STN was identified in Streptomyces flocculus CGMCC 4.1223, revealing an α/β-hydrolase (StnA) and a methyltransferase (StnQ2). In this work, a double mutant ΔstnA/Q2 was constructed by genetic manipulation and produced a novel derivative of STN, named as streptonigramide. Structure of streptonigramide was established by spectroscopic analyses. Its biosynthetic pathway has been proposed as well.
- Published
- 2020
- Full Text
- View/download PDF
19. Tryptophan-Derived Microbial Alkaloids
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Shuangjun Lin, Wenli Guo, Tingting Huang, and Xiaozheng Wang
- Subjects
chemistry.chemical_classification ,endocrine system ,Natural product ,biology ,Drug discovery ,organic chemicals ,Alkaloid ,Tryptophan ,biology.organism_classification ,complex mixtures ,Amino acid ,Actinobacteria ,chemistry.chemical_compound ,chemistry ,Biosynthesis ,Biochemistry ,Nitrogen atom ,heterocyclic compounds - Abstract
Alkaloid natural products comprise a large family of biologically active compounds and many of them exhibit great potential use for biotechnological, agricultural and pharmaceutical industry. Alkaloids are commonly classified according to the amino acid building blocks that provide both the nitrogen atom and the fundamental portion of the skeleton. The first alkaloid was identified in plants, later then, more alkaloids with promising scaffolds for drug discovery and development are of microbial origin, and in particular from filamentous fungi and actinobacteria. We therefore consider alkaloids from microorganisms to be an appropriate starting point to discuss the natural product biosyntheses from the structure-function aspects. This article highlights the microbial alkaloid natural products, including bioactivities, structural diversities, and biosynthesis. The emphasis is on diverse biosynthetic precursors, pathways and novel enzymatic mechanisms. The perspective of strategies for developing structural diversities and creating new structural derivatives will be also discussed.
- Published
- 2020
20. Substrate-bound structures of a ketoreductase from amphotericin modular polyketide synthase
- Author
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Meijuan Yuan, Jianting Zheng, Shuangjun Lin, Xu Xu, Chenguang Liu, Lei Wang, Zixin Deng, and Adrian T. Keatinge-Clay
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chemistry.chemical_classification ,Pantetheine ,Molecular Structure ,biology ,010405 organic chemistry ,Stereochemistry ,Substituent ,Active site ,Stereoisomerism ,010402 general chemistry ,01 natural sciences ,Article ,0104 chemical sciences ,Kinetics ,chemistry.chemical_compound ,Polyketide ,Stereospecificity ,chemistry ,Structural Biology ,Oxidoreductase ,Amphotericin B ,Polyketide synthase ,biology.protein ,Polyketide Synthases ,Ternary complex - Abstract
Ketoreductase (KR) domains of modular polyketide synthases (PKSs) control the stereochemistry of C2 methyl and C3 hydroxyl substituents of polyketide intermediates. To understand the molecular basis of stereocontrol exerted by KRs, the crystal structure of a KR from the second module of the amphotericin PKS (AmpKR2) complexed with NADP+ and 2-methyl-3-oxopentanoyl-pantetheine was solved. This first ternary structure provides direct evidence to the hypothesis that a substrate enters into the active site of an A-type KR from the side opposite the coenzyme to generate an L-hydroxyl substituent. A comparison with the ternary complex of a G355T/Q364H mutant sheds light on the structural basis for stereospecificity toward the substrate C2 methyl substituent. Functional assays suggest the pantetheine handle shows obvious influence on the catalytic efficiency and the stereochemical outcome. Together, these findings extend our current understanding of the stereochemical control of PKS KR domains.
- Published
- 2018
21. Structural basis of the mechanism of β-methyl epimerization by enzyme MarH
- Author
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Yan Hou, Rundong Zhang, Tao Huang, Xiaozheng Wang, Yanli Chen, Shiqi Fang, Xiaofang Ma, Bin Liu, Kaifeng Hu, Shuangjun Lin, and Zhiqiang Bai
- Subjects
0301 basic medicine ,Steric effects ,Models, Molecular ,Indoles ,Chemistry ,Stereochemistry ,Protein Conformation ,Organic Chemistry ,Racemases and Epimerases ,Substrate (chemistry) ,Isomerase ,010402 general chemistry ,01 natural sciences ,Biochemistry ,0104 chemical sciences ,03 medical and health sciences ,030104 developmental biology ,Stereospecificity ,Protein structure ,Molecule ,Physical and Theoretical Chemistry ,Pyruvates ,Isomerization ,Nuclear Magnetic Resonance, Biomolecular ,Carbanion - Abstract
Diverse derivatives of amino acids with different steric configurations are important biosynthetic building blocks. In biology, epimerization is an important way to generate steric diversity. MarH catalyzes the epimerization of the β-position of (3R)-β-methyl-indolepyruvate (MeInPy), forming (3S)-β-MeInPy. Both compounds are derivatives of l-tryptophan (l-Trp) and are important precursors of bioactive natural products. Here, we report the crystal structures of MarH and the NMR structure of its complex with l-Trp, an analogue of its native substrate, (3R)-β-MeInPy. Structural analysis and mutagenesis studies indicated that His25 acts as a base to remove Hβ and generate a planar carbanion intermediate, which is then putatively reprotonated on the opposite face by a water molecule to form (3S)-β-MeInPy in a stereospecific manner. The details of β-site isomerization at the atomic level provide deeper insights into the epimerization mechanism of MarH and will facilitate further enzyme design to extend the substrate scope.
- Published
- 2019
22. Synthesis, antimycobacterial activity and influence on mycobacterial InhA and PknB of 12-membered cyclodepsipeptides
- Author
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Katja Laqua, Linda Liebe, Melissa Richard-Greenblatt, Esther Pérez-Herrán, Ana Guardia, Tingting Huang, Adrian Richter, Peter Imming, Shuangjun Lin, Marcel Klemm, Yossef Av-Gay, and Lluis Ballell
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0301 basic medicine ,Cell Survival ,medicine.drug_class ,Mycobacterium smegmatis ,Clinical Biochemistry ,Antitubercular Agents ,Pharmaceutical Science ,Microbial Sensitivity Tests ,Gram-Positive Bacteria ,Antimycobacterial ,01 natural sciences ,Biochemistry ,Mycobacterium tuberculosis ,03 medical and health sciences ,chemistry.chemical_compound ,Bacterial Proteins ,Cell Line, Tumor ,Depsipeptides ,Gram-Negative Bacteria ,Drug Discovery ,Human Umbilical Vein Endothelial Cells ,medicine ,Humans ,Molecular Biology ,Mycobacterium bovis ,biology ,010405 organic chemistry ,Chemistry ,INHA ,Organic Chemistry ,Isoniazid ,Autophosphorylation ,biology.organism_classification ,0104 chemical sciences ,030104 developmental biology ,Molecular Medicine ,Growth inhibition ,Oxidoreductases ,Proto-Oncogene Proteins c-akt ,medicine.drug - Abstract
In recent years, several small natural cyclopeptides and cyclodepsipeptides were reported to have antimycobacterial activity. Following this lead, a synthetic pathway was developed for a small series of 12-membered ring compounds with one amide and two ester bonds (cyclotridepsipeptides). Within the series, the ring system proved to be necessary for growth inhibition of Mycobacterium smegmatis and Mycobacterium tuberculosis in the low micromolar range. Open-chain precursors and analogues were inactive. The compounds modulated autophosphorylation of the mycobacterial protein kinase B (PknB). PknB inhibitors were active at µM concentration against mycobacteria while inducers were inactive. PknB regulates the activity of the mycobacterial reductase InhA, the target of isoniazid. The activity of the series against Mycobacterium bovis BCG InhA overexpressing strains was indistinguishable from that of the parental strain suggesting that they do not inhibit InhA. All substances were not cytotoxic (HeLa > 5 µg/ml) and did not show any significant antiproliferative effect (HUVEC > 5 µg/ml; K-562 > 5 µg/ml). Within the scope of this study, the molecular target of this new type of small cyclodepsipeptide was not identified, but the data suggest interaction with PknB or other kinases may partly cause the activity.
- Published
- 2018
23. Intramolecular chaperone-mediated secretion of an Rhs effector toxin by a type VI secretion system
- Author
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Tong-Tong Pei, Li-Li Wu, Ming Yu, Guangfeng Liu, Tao G. Dong, Haeun Kim, Zeng-Hang Wang, Hao Li, Ping Xu, Shuangjun Lin, Zhiping Xie, and Xiaoye Liang
- Subjects
0301 basic medicine ,Proteases ,Bacterial toxins ,Operon ,Science ,030106 microbiology ,General Physics and Astronomy ,medicine.disease_cause ,General Biochemistry, Genetics and Molecular Biology ,03 medical and health sciences ,Endonuclease ,Bacterial secretion ,Bacterial Proteins ,medicine ,Pseudomonas syringae ,Secretion ,lcsh:Science ,Bacterial Secretion Systems ,Multidisciplinary ,biology ,Chemistry ,Toxin ,Effector ,General Chemistry ,Gene Expression Regulation, Bacterial ,Type VI Secretion Systems ,Publisher Correction ,Cell biology ,030104 developmental biology ,Genes, Bacterial ,Chaperone (protein) ,Mutation ,Pseudomonas aeruginosa ,biology.protein ,lcsh:Q ,Aeromonas ,Pathogens ,Molecular Chaperones ,Peptide Hydrolases - Abstract
Bacterial Rhs proteins containing toxic domains are often secreted by type VI secretion systems (T6SSs) through unclear mechanisms. Here, we show that the T6SS Rhs-family effector TseI of Aeromonas dhakensis is subject to self-cleavage at both the N- and the C-terminus, releasing the middle Rhs core and two VgrG-interacting domains (which we name VIRN and VIRC). VIRC is an endonuclease, and the immunity protein TsiI protects against VIRC toxicity through direct interaction. Proteolytic release of VIRC and VIRN is mediated, respectively, by an internal aspartic protease activity and by two conserved glutamic residues in the Rhs core. Mutations abolishing self-cleavage do not block secretion, but reduce TseI toxicity. Deletion of VIRN or the Rhs core abolishes secretion. TseI homologs from Pseudomonas syringae, P. aeruginosa, and Vibrio parahaemolyticus are also self-cleaved. VIRN and VIRC interact with protein VgrG1, while the Rhs core interacts with protein TecI. We propose that VIRN and the Rhs core act as T6SS intramolecular chaperones to facilitate toxin secretion and function. Bacterial Rhs proteins with toxic domains are often secreted by type VI secretion systems. Here, the authors show that one of these proteins self-cleaves into three fragments, with the Rhs core and the N-terminal domain facilitating secretion and function of the C-terminal toxic domain.
- Published
- 2019
24. Biosynthetic study of β-methyl amino acid building blocks involved in natural products
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ShuangJun Lin and YuYang Zhang
- Subjects
chemistry.chemical_classification ,Multidisciplinary ,Stereochemistry ,Primary metabolite ,Biology ,010502 geochemistry & geophysics ,01 natural sciences ,Amino acid ,Metabolic engineering ,chemistry.chemical_compound ,Protein catabolism ,Enzyme ,Biosynthesis ,chemistry ,Biochemistry ,Aspartic acid ,Radical SAM ,0105 earth and related environmental sciences - Abstract
Natural products are an important source of pharmaceuticals or drug leads for the human lifes. The biosynthetic studies on these small chemical molecules can offer the possibilities to improve the titers and produce a lot of non-natural natural products with enhanced biological activity. Amino acids are a type of important building blocks of natural products, and apart from twenty proteinogenic amino acids, about five hundred nonproteinogenic amino acids were found to be involved into the biosynthesis of natural products. These nonproteinogenic amino acid units in natural products have been confirmed to play crucial roles in their stabilities and biological activities. β-methyl amino acid is one kind of nonproteinogenic amino acids and has been found in many bioactive secondary metabolites, including daptomycin, streptonigrin, indolemycin and so on, and as well as several primary metabolites such as 3-methyl-glutamate in fermentation process of several Clostridium sp. The β-carbon of amino acids is not an active position for methylation, therefore the methylation at the β-position is still chemically very challenging. Recently, the biosynthetic studies of microbial natural products revealed many novel enzymatic reactions or pathways for amino acid modifications including the β-methylation of amino acids. Up to now, three different biosynthetic pathways for β-methyl amino acids have been reported, including B12-dependent glutamate aminomutase, combination of a methyltransferase and an aminotranferase, and B12-dependent radical SAM methyltransferase. B12-dependent glutamate aminomutase catalyzes the rearrangement of glutamate to generate β-methyl aspartic acid, a building black of Nikkomycins and other natural products. Combination of a methyltransferase and an aminotranferase uses the aminotransferase to generate α-keto acids as the active substrates for the methylation catalzyed by the methyltransferase from amino acids while B12-dependent radical SAM methyltransferase can catalyze the direct methylation at the β-carbon of amino acids via a radical manner. There have been many reviews that summarized the enzymatic modifications of amino acids, but there has been none of literatures covering this type of intereting and useful nonproteinogenic amino acids. This review summarized the recent advances in the biosynthesis of β-methyl amino acids as the building blocks of several natural products and discussed the biosynthetic or enzymatic mechanisms involved in the β-methylation of amino acids in detail. Basesd on the mechanisms of the biosynthesis of these β-methyl amino acids, the application of β-methyl amino acids was proposed in this review. This study may provide new strategies and useful information for improving the yields of the valuable natural products containing β-methyl amino acids and introducing these building blocks into more chemical scaffolds by metabolic engineering, synthetic biology and enzyme engineering methods.
- Published
- 2017
25. Biosynthesis of the pyrrolidine protein synthesis inhibitor anisomycin involves novel gene ensemble and cryptic biosynthetic steps
- Author
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Shuangjun Lin, Bo Cao, Xiaoqing Zheng, Xiaozheng Wang, Zixin Deng, Yit-Heng Chooi, Delin You, Lingxin Kong, Min Xu, Qiuxiang Cheng, and Fen Yao
- Subjects
0301 basic medicine ,Glycosylation ,Bacterial genome size ,Streptomyces ,Pyrrolidine ,03 medical and health sciences ,chemistry.chemical_compound ,Biosynthesis ,Gene cluster ,Anisomycin ,Multidisciplinary ,biology ,Computational Biology ,Biological Sciences ,biology.organism_classification ,Reverse genetics ,Anti-Bacterial Agents ,Biosynthetic Pathways ,High-Throughput Screening Assays ,030104 developmental biology ,chemistry ,Biochemistry ,Multigene Family ,Genome, Bacterial - Abstract
The protein synthesis inhibitor anisomycin features a unique benzylpyrrolidine system and exhibits diverse biological and pharmacologic activities. Its biosynthetic origin has remained obscure for more than 60 y, however. Here we report the identification of the biosynthetic gene cluster (BGC) of anisomycin in Streptomyces hygrospinosus var. beijingensis by a bioactivity-guided high-throughput screening method. Using a combination of bioinformatic analysis, reverse genetics, chemical analysis, and in vitro biochemical assays, we have identified a core four-gene ensemble responsible for the synthesis of the pyrrolidine system in anisomycin: aniQ, encoding a aminotransferase that catalyzes an initial deamination and a later reamination steps; aniP, encoding a transketolase implicated to bring together an glycolysis intermediate with 4-hydroxyphenylpyruvic acid to form the anisomycin molecular backbone; aniO, encoding a glycosyltransferase that catalyzes a cryptic glycosylation crucial for downstream enzyme processing; and aniN, encoding a bifunctional dehydrogenase that mediates multistep pyrrolidine formation. The results reveal a BGC for pyrrolidine alkaloid biosynthesis that is distinct from known bacterial alkaloid pathways, and provide the signature sequences that will facilitate the discovery of BGCs encoding novel pyrrolidine alkaloids in bacterial genomes. The biosynthetic insights from this study further set the foundation for biosynthetic engineering of pyrrolidine antibiotics.
- Published
- 2017
26. Characterization of an 17β-estradiol-degrading bacterium Stenotrophomonas maltophilia SJTL3 tolerant to adverse environmental factors
- Author
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Weiliang Xiong, Wanli Peng, Zixin Deng, Rubing Liang, Shuangjun Lin, and Chong Yin
- Subjects
Microorganism ,Stenotrophomonas maltophilia ,Estrone ,Applied Microbiology and Biotechnology ,03 medical and health sciences ,chemistry.chemical_compound ,Bioremediation ,Stress, Physiological ,Metals, Heavy ,Soil Pollutants ,Food science ,Phylogeny ,030304 developmental biology ,0303 health sciences ,Microbial Viability ,biology ,Strain (chemistry) ,Estradiol ,Sewage ,030306 microbiology ,Estrogens ,General Medicine ,Contamination ,Biodegradation ,Hydrogen-Ion Concentration ,biology.organism_classification ,Kinetics ,Biodegradation, Environmental ,chemistry ,Bacteria ,Genome, Bacterial ,Biotechnology - Abstract
Bioremediation of environmental estrogens requires microorganisms with stable degradation efficiency and great stress tolerance in complex environments. In this work, Stenotrophomonas maltophilia SJTL3 isolated from wastewater was found to be able to degrade over 90% of 10 μg/mL 17β-estradiol (E2) in a week and the degradation dynamic was fitted by the first-order kinetic equations. Estrone was the first and major intermediate of E2 biodegradation. Strain SJTL3 exhibited strong tolerance to several adverse conditions like extreme pH (3.0–11.0), high osmolality (2%), co-existing heavy metals (6.25 μg/mL of Cu2+) and surfactants (5 CMC of Tween 80), and retained normal cell vitality and stable E2-degradaing efficiency. In solid soil, strain SJTL3 could remove nearly 100% of 1 μg/mL of E2 after the bacteria inoculation and 8-day culture. As to the contamination of 10 μg/mL E2 in soil, the biodegradation efficiency was about 90%. The further obtainment of the whole genome of strain SJTL3 and genome analysis revealed that this strain contained not only the potential genes responsible for estrogen degradation, but also the genes encoding proteins involved in stress tolerance. This work could promote the estrogen-biodegrading mechanism study and provide insights into the bioremediation application.
- Published
- 2019
27. Characterization of an efficient estrogen-degrading bacterium Stenotrophomonas maltophilia SJTH1 in saline-, alkaline-, heavy metal-contained environments or solid soil and identification of four 17β-estradiol-oxidizing dehydrogenases
- Author
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Chong Yin, Yanqiu Wang, Weiliang Xiong, Rubing Liang, Shuangjun Lin, and Zixin Deng
- Subjects
Bioaugmentation ,Environmental Engineering ,Octoxynol ,Health, Toxicology and Mutagenesis ,Microorganism ,Stenotrophomonas maltophilia ,0211 other engineering and technologies ,Polysorbates ,02 engineering and technology ,010501 environmental sciences ,01 natural sciences ,Estradiol Dehydrogenases ,Surface-Active Agents ,Bioremediation ,Bacterial Proteins ,Environmental Chemistry ,Food science ,Amino Acid Sequence ,Waste Management and Disposal ,0105 earth and related environmental sciences ,021110 strategic, defence & security studies ,Strain (chemistry) ,biology ,Estradiol ,Chemistry ,Biodegradation ,Contamination ,biology.organism_classification ,Pollution ,Kinetics ,Biodegradation, Environmental ,Oxidation-Reduction ,Sequence Alignment ,Bacteria - Abstract
The efficient bioremediation of estrogen contamination in complex environments is of great concern. Here the strain Stenotrophomonas maltophilia SJTH1 was found with great and stable estrogen-degradation efficiency even under stress environments. The strain could utilize 17β-estradiol (E2) as a carbon source and degrade 90% of 10 mg/L E2 in a week; estrone (E1) was the first degrading intermediate of E2. Notably, diverse pH conditions (3.0–11.0) and supplements of 4% salinity, 6.25 mg/L of heavy metal (Cd2+ or Cu2+), or 1 CMC of surfactant (Tween 80/ Triton X-100) had little effect on its cell growth and estrogen degradation. The addition of low concentrations of copper and Tween 80 even promoted its E2 degradation. Bioaugmentation of strain SJTH1 into solid clay soil achieved over 80% removal of E2 contamination (10 mg/kg) within two weeks. Further, the whole genome sequence of S. maltophilia SJTH1 was obtained, and a series of potential genes participating in stress-tolerance and estrogen-degradation were predicted. Four dehydrogenases similar to 17β-hydroxysteroid dehydrogenases (17β-HSDs) were found to be induced by E2, and the four heterogenous-expressed enzymes could oxidize E2 into E1 efficiently. This work could promote bioremediation appliance potential with microorganisms and biodegradation mechanism study of estrogens in complex real environments.
- Published
- 2019
28. Design and Biosynthesis of Dimeric Alboflavusins with Biaryl Linkages via Regiospecific C-C Bond Coupling
- Author
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Zhengyan Guo, Chao Li, Shuangjun Lin, Guozhu Chen, Pengwei Li, Hua Xiang, Jianhua Ju, Yihua Chen, Zhenju Cao, Jinwei Ren, and Yuwei Zhang
- Subjects
0301 basic medicine ,chemistry.chemical_classification ,Natural product ,Streptomyces alboflavus ,010405 organic chemistry ,Stereochemistry ,Chemistry ,Mutant ,General Chemistry ,01 natural sciences ,Biochemistry ,Catalysis ,In vitro ,0104 chemical sciences ,03 medical and health sciences ,chemistry.chemical_compound ,030104 developmental biology ,Colloid and Surface Chemistry ,Biosynthesis ,Nonribosomal peptide ,Gene cluster ,Gene - Abstract
Alboflavusins (AFNs) are a group of cyclohexapeptides with moderate antibacterial and antitumor activities from Streptomyces alboflavus sp. 313. In vivo and in vitro studies proposed that AFNs are biosynthesized by a nonribosomal peptide synthetase machinery, and the 6-Cl-L-Trp precursor is supplied by a tryptophan halogenase gene located outside the afn gene cluster. Guided by the structure-activity relationship knowledge about the AFN-like cyclohexapeptides, two dimeric AFNs (di-AFNs) with regiospecific biaryl linkages were designed and generated biotechnologically by expressing the P450 gene hmtS or clpS in S. alboflavus wild-type and mutant strains. The di-AFNs displayed much better antibacterial and antitumor activities than their monomers as anticipated, exemplifying a rational strategy to generate natural product congeners with improved bioactivities.
- Published
- 2018
29. Indole methylation protects diketopiperazine configuration in the maremycin biosynthetic pathway
- Author
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Shuangjun Lin, Zixin Deng, Xiaozheng Wang, Nelson L. Brock, Yingxia Lan, Yi Zou, and Tingting Huang
- Subjects
Indole test ,Methyltransferase ,biology ,010405 organic chemistry ,Chemistry ,Stereochemistry ,Mutant ,Tryptophan ,General Chemistry ,Methylation ,010402 general chemistry ,biology.organism_classification ,01 natural sciences ,Streptomyces ,0104 chemical sciences ,chemistry.chemical_compound ,Biochemistry ,Biosynthesis ,Gene cluster - Abstract
The maremycin biosynthetic gene cluster has been identified in Streptomyces sp. B9173. Comparative metabolic profiling with knockout mutant strains led to the identification of new products correlated to the maremycin biosynthesis, in particular the “demethyl”-maremycins with an unexpected D-tryptophan unit. A biosynthetic pathway for the maremycins is proposed and plausible reasoning for tryptophan epimerization in the demethylmaremycin biosynthesis is also provided.
- Published
- 2016
30. Transformation of Streptonigrin to Streptonigrone: Flavin Reductase-Mediated Flavin-Catalyzed Concomitant Oxidative Decarboxylation of Picolinic Acid Derivatives
- Author
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Shuangjun Lin, Xiufen Zhou, Fei Xu, Dekun Kong, Zixin Deng, Nelson L. Brock, and Jing Wo
- Subjects
010405 organic chemistry ,Decarboxylation ,Chemistry ,General Chemistry ,Flavin group ,Picolinic acid ,010402 general chemistry ,01 natural sciences ,Combinatorial chemistry ,Catalysis ,0104 chemical sciences ,Hydroxylation ,chemistry.chemical_compound ,Streptonigrin ,Flavin reductase ,Organic chemistry ,Oxidative decarboxylation - Abstract
In the flavin-reductase-catalyzed reducing condition, a mild and efficient α-hydroxylation and decarboxylation procedure using natural flavins as a catalyst and atmospheric oxygen as an external oxidizing agent has been successfully developed and applied to the synthesis of streptonigrone from streptonigrin. This reaction was achieved not only with streptonigrin analogues but also with structurally diverse electron-rich picolinic acid derivatives. The hydroxylation and decarboxylation may take place in a concerted manner.
- Published
- 2016
31. An Acyl Transfer Reaction Catalyzed by an Epimerase MarH
- Author
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Haixing Yin, Yiwen Chu, Tingting Huang, Zixin Deng, Nelson L. Brock, Yi Zou, Shuangjun Lin, and Mo Han
- Subjects
chemistry.chemical_classification ,010405 organic chemistry ,Stereochemistry ,Chemistry ,Mutagenesis ,Substrate (chemistry) ,General Chemistry ,010402 general chemistry ,01 natural sciences ,Catalysis ,0104 chemical sciences ,Amino acid ,Acylation ,Chloramphenicol acetyltransferase ,chemistry.chemical_compound ,Nucleophile ,Biosynthesis ,Acyltransferase ,lipids (amino acids, peptides, and proteins) - Abstract
MarH, a small protein (129 amino acids) belonging to the cupin superfamily, was previously characterized as an epimerase involved in the (2S,3S)-β-methyltryptophan formation in the maremycin biosynthesis. Here, MarH was discovered to act as an acyltransferase that can catalyze the 3-O-acylation of chloramphenicol. Furthermore, MarH can catalyze N-acylation of deacylated chloramphenicol analogue thereby activating them for 3-O-acylation. By systematic site-directed mutagenesis, H64 was revealed as a potential catalytic base that deprotonates the acyl acceptor substrate. Nucleophilic attack at the carbonyl carbon of the acyl donor then gives the acylation product.
- Published
- 2016
32. Production of high-value drug precursors by the whole-cell catalyst based on the transformation of ring-hydroxylating dioxygenase to aromatic compounds
- Author
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Zixin Deng, Shuangjun Lin, Rubing Liang, Weiliang Xiong, Mancheng Tang, Lin Chen, Hua Qiu, and Chong Yin
- Subjects
Environmental Engineering ,biology ,Renewable Energy, Sustainability and the Environment ,Chemistry ,020209 energy ,Bioengineering ,Hydrochloric acid ,02 engineering and technology ,010501 environmental sciences ,Ring (chemistry) ,biology.organism_classification ,01 natural sciences ,Chemical synthesis ,Sphingobium ,Catalysis ,Dibenzofuran ,chemistry.chemical_compound ,Dioxygenase ,Yield (chemistry) ,0202 electrical engineering, electronic engineering, information engineering ,Organic chemistry ,Waste Management and Disposal ,0105 earth and related environmental sciences - Abstract
Monohydroxy heterocyclic aromatic compounds are the widely-used precursors for pharmaceutical synthesis. In this work a whole-cell catalyst was constructed to generate hydroxylated aromatics efficiently using the ring-hydroxylating dioxygenase (RHD) from Sphingobium yanoikuyae SJTF8. With the whole-cell catalyst and 10-min hydrochloric acid treatment, dibenzofuran was transformed into 2-hydroxydibenzofuran with the yield of 90.7% in 6 h and the chemical value was enhanced 74 times. Various monohydroxy aromatics or dihydroxy aromatics were also generated from the typical and heterocyclic aromatics waste by the whole-cell catalyst. This whole-cell catalyst provides a simple, economic, green and efficient strategy to produce the value-added aromatic drug precursors and presents a great supplement of chemical synthesis.
- Published
- 2020
33. Publisher Correction: Intramolecular chaperone-mediated secretion of an Rhs effector toxin by a type VI secretion system
- Author
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Shuangjun Lin, Ping Xu, Tao G. Dong, Tong-Tong Pei, Guangfeng Liu, Zhiping Xie, Li-Li Wu, Xiaoye Liang, Hao Li, Ming Yu, Zeng-Hang Wang, and Haeun Kim
- Subjects
Multidisciplinary ,biology ,Bacterial toxins ,Chemistry ,Effector ,Toxin ,Science ,General Physics and Astronomy ,General Chemistry ,Proteases ,medicine.disease_cause ,General Biochemistry, Genetics and Molecular Biology ,Article ,Cell biology ,Bacterial secretion ,Chaperone (protein) ,Intramolecular force ,biology.protein ,medicine ,Secretion ,lcsh:Q ,Pathogens ,lcsh:Science - Abstract
Bacterial Rhs proteins containing toxic domains are often secreted by type VI secretion systems (T6SSs) through unclear mechanisms. Here, we show that the T6SS Rhs-family effector TseI of Aeromonas dhakensis is subject to self-cleavage at both the N- and the C-terminus, releasing the middle Rhs core and two VgrG-interacting domains (which we name VIRN and VIRC). VIRC is an endonuclease, and the immunity protein TsiI protects against VIRC toxicity through direct interaction. Proteolytic release of VIRC and VIRN is mediated, respectively, by an internal aspartic protease activity and by two conserved glutamic residues in the Rhs core. Mutations abolishing self-cleavage do not block secretion, but reduce TseI toxicity. Deletion of VIRN or the Rhs core abolishes secretion. TseI homologs from Pseudomonas syringae, P. aeruginosa, and Vibrio parahaemolyticus are also self-cleaved. VIRN and VIRC interact with protein VgrG1, while the Rhs core interacts with protein TecI. We propose that VIRN and the Rhs core act as T6SS intramolecular chaperones to facilitate toxin secretion and function., Bacterial Rhs proteins with toxic domains are often secreted by type VI secretion systems. Here, the authors show that one of these proteins self-cleaves into three fragments, with the Rhs core and the N-terminal domain facilitating secretion and function of the C-terminal toxic domain.
- Published
- 2020
34. NRPS Protein MarQ Catalyzes Flexible Adenylation and Specific S-Methylation
- Author
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Shuangjun Lin, Zixin Deng, Yi Zou, Yingyi Duan, and Tingting Huang
- Subjects
Methyltransferase ,Indoles ,Stereochemistry ,010402 general chemistry ,01 natural sciences ,Biochemistry ,Methylation ,Reaction sequence ,Bacterial Proteins ,Nonribosomal peptide ,Humans ,Peptidyl carrier protein ,Cysteine ,Peptide Synthases ,Adenylylation ,Piperazine ,chemistry.chemical_classification ,010405 organic chemistry ,Chemistry ,General Medicine ,Streptomyces ,0104 chemical sciences ,Amino acid ,Biosynthetic Pathways ,Thiol ,Molecular Medicine - Abstract
Maremycins are a group of structurally diverse 2,5-diketopiperazine natural products featuring a rare amino acid building block, S-methyl-l-cysteine (Me-Cys). Three freestanding nonribosomal peptide synthetase (NRPS) proteins from the maremycins biosynthetic pathway were proposed for the formation of the 2,5-diketopiperazine scaffold: MarQ, MarM, and MarJ. MarQ displays flexible adenylation activity toward Cys, Me-Cys, Ser, and ( S)-2,3-diaminopropanoic acid (DAP) and transfers these substrates to MarJ, which is the discrete peptidyl carrier protein (PCP). MarQ could also activate several other amino acids. The embedded methyltransferase (MT) domain in MarQ specifically catalyzes the thiol methylation of MarJ-tethered Cys. The in vitro reconstitution of MarQ and MarJ further provides clear evidence for the reaction sequence of methylation step on Cys. Our study on MarJ/Q tridomain cassette gains valuable insights into maremycins structure diversity and will be exploited to incorporate Me-Cys into natural products by combinatorial biosynthesis.
- Published
- 2018
35. Divergent biosynthesis of indole alkaloids FR900452 and spiro-maremycins
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Tingting Huang, Kaifeng Hu, Zixin Deng, Tao Huang, Yanyan Liu, Shuangjun Lin, Yi Zou, and Yingyi Duan
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0301 basic medicine ,Cyclopentenone ,Indoles ,Stereochemistry ,Cyclopentanes ,Diketopiperazines ,010402 general chemistry ,01 natural sciences ,Biochemistry ,Streptomyces ,Indole Alkaloids ,03 medical and health sciences ,chemistry.chemical_compound ,Biosynthesis ,Gene cluster ,Moiety ,Spiro Compounds ,Physical and Theoretical Chemistry ,Gene ,Indole test ,biology ,Chemistry ,Organic Chemistry ,biology.organism_classification ,0104 chemical sciences ,Biosynthetic Pathways ,030104 developmental biology ,Multigene Family ,Fermentation ,Heterologous expression - Abstract
FR900452 and spirocyclic maremycins, including F and G components, are structurally related indole alkaloids, previously identified from different Streptomyces species. These alkaloids feature an indole diketopiperazine motif linked with a cyclopentenone moiety, but the linkage differs in FR900452 and the spirocyclic maremycins. Here, FR900452 and its two new analogues were identified from the fermentation broth of Streptomyces sp. B9173, the producer of maremycins. Gene inactivation and heterologous expression of the mar gene cluster confirmed that production of FR900452 shares the same biosynthetic machinery that produces maremycins. FR900452 was identified as the precursor of maremycin A/B by feeding studies. MarP, a SnoaL-like protein, was demonstrated to differentiate the biosynthesis of FR900452 from that of spiro-form maremycin G.
- Published
- 2018
36. Biosynthesis of Tropolones in Streptomyces spp.: Interweaving Biosynthesis and Degradation of Phenylacetic Acid and Hydroxylations on the Tropone Ring
- Author
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Meifeng Tao, Zixin Deng, Jianguo Xu, Shuangjun Lin, Xuefei Chen, Min Xu, Yemin Wang, and Jin Lü
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0301 basic medicine ,Genetics and Molecular Biology ,Prephenate dehydratase ,Phenylacetic acid ,Hydroxylation ,Applied Microbiology and Biotechnology ,Streptomyces ,Tropolone ,Metabolic engineering ,03 medical and health sciences ,chemistry.chemical_compound ,Bacterial Proteins ,Biosynthesis ,Gene cluster ,Phenylacetates ,Molecular Structure ,Ecology ,biology ,biology.organism_classification ,Biosynthetic Pathways ,030104 developmental biology ,chemistry ,Biochemistry ,Multigene Family ,Chorismate mutase ,Gene Deletion ,Food Science ,Biotechnology - Abstract
Tropolonoids are important natural products that contain a unique seven-membered aromatic tropolone core and exhibit remarkable biological activities. 3,7-Dihydroxytropolone (DHT) isolated from Streptomyces species is a multiply hydroxylated tropolone exhibiting antimicrobial, anticancer, and antiviral activities. In this study, we determined the DHT biosynthetic pathway by heterologous expression, gene deletion, and biotransformation. Nine trl genes and some of the aerobic phenylacetic acid degradation pathway genes ( paa ) located outside the trl biosynthetic gene cluster are required for the heterologous production of DHT. The trlA gene encodes a single-domain protein homologous to the C-terminal enoyl coenzyme A (enoyl-CoA) hydratase domain of PaaZ. TrlA truncates the phenylacetic acid catabolic pathway and redirects it toward the formation of heptacyclic intermediates. TrlB is a 3-deoxy- d -arabino-heptulosonic acid-7-phosphate (DAHP) synthase homolog. TrlH is an unusual bifunctional protein bearing an N-terminal prephenate dehydratase domain and a C-terminal chorismate mutase domain. TrlB and TrlH enhanced de novo biosynthesis of phenylpyruvate, thereby providing abundant precursor for the prolific production of DHT in Streptomyces spp. Six seven-membered carbocyclic compounds were identified from the trlC , trlD , trlE , and trlF deletion mutants. Four of these chemicals, including 1,4,6-cycloheptatriene-1-carboxylic acid, tropone, tropolone, and 7-hydroxytropolone, were verified as key biosynthetic intermediates. TrlF is required for the conversion of 1,4,6-cycloheptatriene-1-carboxylic acid into tropone. The monooxygenases TrlE and TrlCD catalyze the regioselective hydroxylations of tropone to produce DHT. This study reveals a natural association of anabolism of chorismate and phenylpyruvate, catabolism of phenylacetic acid, and biosynthesis of tropolones in Streptomyces spp. IMPORTANCE Tropolonoids are promising drug lead compounds because of the versatile bioactivities attributed to their highly oxidized seven-membered aromatic ring scaffolds. Our present study provides clear insight into the biosynthesis of 3,7-dihydroxytropolone (DHT) through the identification of key genes responsible for the formation and modification of the seven-membered aromatic core. We also reveal the intrinsic mechanism of elevated production of DHT and related tropolonoids in Streptomyces spp. The study on DHT biosynthesis in Streptomyces exhibits a good example of antibiotic production in which both anabolic and catabolic pathways of primary metabolism are interwoven into the biosynthesis of secondary metabolites. Furthermore, our study sets the stage for metabolic engineering of the biosynthetic pathway for natural tropolonoid products and provides alternative synthetic biology tools for engineering novel tropolonoids.
- Published
- 2018
37. Conserved residues Arg188 and Asp302 are critical for active site organization and catalysis in human ABO(H) blood group A and B glycosyltransferases
- Author
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Svetlana N. Borisova, Brock Schuman, Ruixiang Blake Zheng, Shuangjun Lin, Mattias Persson, Omid Haji-Ghassemi, Susannah M. L. Gagnon, James A. Letts, Robert Polakowski, Monica M. Palcic, Stephen V. Evans, Max S G Legg, and Brian P. Rempel
- Subjects
0301 basic medicine ,Stereochemistry ,Protein domain ,Mutant ,Disaccharide ,H antigen ,Arginine ,Crystallography, X-Ray ,Biochemistry ,ABO Blood-Group System ,Regular Manuscripts ,03 medical and health sciences ,chemistry.chemical_compound ,Protein Domains ,Catalytic Domain ,Glycosyltransferase ,Transferase ,Humans ,chemistry.chemical_classification ,Aspartic Acid ,030102 biochemistry & molecular biology ,biology ,Active site ,Glycosyltransferases ,030104 developmental biology ,Enzyme ,chemistry ,biology.protein - Abstract
Homologous glycosyltransferases GTA and GTB perform the final step in human ABO(H) blood group A and B antigen synthesis by transferring the sugar moiety from donor UDP-GalNAc/UDP-Gal to the terminal H antigen disaccharide acceptor. Like other GT-A fold family 6 glycosyltransferases, GTA and GTB undergo major conformational changes in two mobile regions, the C-terminal tail and internal loop, to achieve the closed, catalytic state. These changes are known to establish a salt bridge network among conserved active site residues Arg188, Asp211 and Asp302, which move to accommodate a series of discrete donor conformations while promoting loop ordering and formation of the closed enzyme state. However, the individual significance of these residues in linking these processes remains unclear. Here, we report the kinetics and high-resolution structures of GTA/GTB mutants of residues 188 and 302. The structural data support a conserved salt bridge network critical to mobile polypeptide loop organization and stabilization of the catalytically competent donor conformation. Consistent with the X-ray crystal structures, the kinetic data suggest that disruption of this salt bridge network has a destabilizing effect on the transition state, emphasizing the importance of Arg188 and Asp302 in the glycosyltransfer reaction mechanism. The salt bridge network observed in GTA/GTB structures during substrate binding appears to be conserved not only among other Carbohydrate Active EnZyme family 6 glycosyltransferases but also within both retaining and inverting GT-A fold glycosyltransferases. Our findings augment recently published crystal structures, which have identified a correlation between donor substrate conformational changes and mobile loop ordering.
- Published
- 2018
38. Operon for Biosynthesis of Lipstatin, the Beta-Lactone Inhibitor of Human Pancreatic Lipase
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Qingshan Long, Tingli Bai, Yemin Wang, Meifeng Tao, Wen Liu, Hong-Yu Ou, Zixin Deng, Daozhong Zhang, Qianjin Kang, and Shuangjun Lin
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Linoleic acid ,Genetics and Molecular Biology ,Lipstatin ,Applied Microbiology and Biotechnology ,Lactones ,chemistry.chemical_compound ,Bacterial Proteins ,Biosynthesis ,Nonribosomal peptide ,Operon ,Humans ,Enzyme Inhibitors ,Lipase ,chemistry.chemical_classification ,Ecology ,biology ,Streptomyces toxytricini ,Fatty acid ,biology.organism_classification ,Streptomyces ,Biosynthetic Pathways ,chemistry ,Biochemistry ,biology.protein ,Leucine ,Food Science ,Biotechnology - Abstract
Lipstatin, isolated from Streptomyces toxytricini as a potent and selective inhibitor of human pancreatic lipase, is a precursor for tetrahydrolipstatin (also known as orlistat, Xenical, and Alli), the only FDA-approved antiobesity medication for long-term use. Lipstatin features a 2-hexyl-3,5-dihydroxy-7,10-hexadecadienoic-β-lactone structure with an N -formyl- l -leucine group attached as an ester to the 5-hydroxy group. It has been suggested that the α-branched 3,5-dihydroxy fatty acid β-lactone moiety of lipstatin in S. toxytricini is derived from Claisen condensation between two fatty acid substrates, which are derived from incomplete oxidative degradation of linoleic acid based on feeding experiments. In this study, we identified a six-gene operon ( lst ) that was essential for the biosynthesis of lipstatin by large-deletion, complementation, and single-gene knockout experiments. lstA , lstB , and lstC , which encode two β-ketoacyl–acyl carrier protein synthase III homologues and an acyl coenzyme A (acyl-CoA) synthetase homologue, were indicated to be responsible for the generation of the α-branched 3,5-dihydroxy fatty acid backbone. Subsequently, the nonribosomal peptide synthetase (NRPS) gene lstE and the putative formyltransferase gene lstF were involved in decoration of the α-branched 3,5-dihydroxy fatty acid chain with an N-formylated leucine residue. Finally, the 3β-hydroxysteroid dehydrogenase-homologous gene lstD might be responsible for the reduction of the β-keto group of the biosynthetic intermediate, thereby facilitating the formation of the unique β-lactone ring.
- Published
- 2014
39. Formation of an Angular Aromatic Polyketide from a Linear Anthrene Precursor via Oxidative Rearrangement
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Zixin Deng, Ying Wang, Meifeng Tao, Jürgen Rohr, Min Xu, Hong-Yu Ou, Lijun Xu, Ming Jiang, Fei Zhang, Guixi Gao, Shuangjun Lin, Jin Xiong Lv, Yemin Wang, Xiangyang Liu, Qianjin Kang, and Qingshan Long
- Subjects
0301 basic medicine ,Magnetic Resonance Spectroscopy ,Stereochemistry ,Clinical Biochemistry ,Oxidative phosphorylation ,010402 general chemistry ,01 natural sciences ,Biochemistry ,Mass Spectrometry ,Article ,03 medical and health sciences ,Polyketide ,chemistry.chemical_compound ,Biosynthesis ,Oxidoreductase ,Drug Discovery ,Molecular Biology ,Pharmacology ,chemistry.chemical_classification ,Anthracenes ,Polyketide formation ,Quinones ,Phenanthrenes ,Streptomyces ,0104 chemical sciences ,Folding (chemistry) ,030104 developmental biology ,chemistry ,Multigene Family ,Polyketides ,Biocatalysis ,Molecular Medicine ,Streptomyces lividans ,Oxidoreductases ,Polyketide Synthases - Abstract
Summary Bacterial aromatic polyketides are a group of natural products synthesized by polyketide synthases (PKSs) that show diverse structures and biological activities. They are structurally subclassified into linear, angular, and discoid aromatic polyketides, the formation of which is commonly determined by the shaping and folding of the poly-β-keto intermediates under the concerted actions of the minimal PKSs, cyclases and ketoreductases. Murayaquinone, found in several streptomycetes, possesses an unusual tricyclic angular aromatic polyketide core containing a 9,10-phenanthraquinone. In this study, genes essential for murayaquinone biosynthesis were identified, and a linear anthraoxirene intermediate was discovered. A unique biosynthetic model for the angular aromatic polyketide formation was discovered and confirmed through in vivo and in vitro studies. Three oxidoreductases, MrqO3, MrqO6, and MrqO7, were identified to catalyze the conversion of the linear aromatic polyketide intermediate into the final angularly arranged framework, which exemplifies a novel strategy for the biosynthesis of angular aromatic polyketides.
- Published
- 2017
40. A new glutarimide derivative from marine sponge-derived Streptomyces anulatus S71
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Zhiyong Li, Dandan Sun, Zixin Deng, Yinxian Yu, Wei Sun, and Shuangjun Lin
- Subjects
Indoles ,Stereochemistry ,Ethyl acetate ,Marine Biology ,Glutarimide ,Plant Science ,Biochemistry ,Analytical Chemistry ,chemistry.chemical_compound ,Polyketide synthase ,Animals ,Hydroxymethyl ,Nuclear Magnetic Resonance, Biomolecular ,Piperidones ,Indole test ,Molecular Structure ,Indole alkaloid ,biology ,Organic Chemistry ,Streptomyces ,Porifera ,chemistry ,biology.protein ,Polyketide Synthases ,Derivative (chemistry) ,Streptomyces anulatus - Abstract
Four glutarimide-derived compounds including a new 3-[2-[2-hydroxy-3-methylphenyl-5-(hydroxymethyl)]-2-oxoethyl] glutarimide (1) and three known 3-[2-(2-hyroxy-3,5- dimethylphenyl)-2-oxoethyl] glutarimide (2, actiphenol), 3-hydroxy-3-[2-(2-hydroxy-3,5-dimethylphenyl)-2-oxoethyl] glutarimide (3) and 3-[2-[2-hydroxy-3-(hydroxymethyl)-5-methylphenyl]-2-oxoethyl] glutarimide (4), along with a known indole alkaloid 3-(hydroxyacetyl) indole (5), were isolated from ethyl acetate extract of the fermentation broth of the marine sponge-derived Streptomyces anulatus S71. Their structures were deduced by extensive studies of NMR and mass spectra.
- Published
- 2014
41. Significance of agitation-induced shear stress on mycelium morphology and lavendamycin production by engineered Streptomyces flocculus
- Author
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Xue Xia, Xiao-Xia Xia, Feng-Song Cong, Jian-Jiang Zhong, and Shuangjun Lin
- Subjects
Antibiotics, Antineoplastic ,Hypha ,Strain (chemistry) ,General Medicine ,equipment and supplies ,Cell morphology ,Applied Microbiology and Biotechnology ,Streptomyces ,Oxygen ,chemistry.chemical_compound ,chemistry ,Botany ,Pellet ,Bioreactor ,Shear stress ,Streptonigrin ,Stress, Mechanical ,Food science ,Lavendamycin ,Mycelium ,Mechanical Phenomena ,Biotechnology - Abstract
Lavendamycin methyl ester (LME) is a derivative of a highly functionalized aminoquinone alkaloid lavendamycin and could be used as a scaffold for novel anticancer agent development. This work demonstrated LME production by cultivation of an engineered strain of Streptomyces flocculus CGMCC4.1223 ΔstnB1, while the wild-type strain did not produce. To enhance its production, the effect of shear stress and oxygen supply on ΔstnB1 strain cultivation was investigated in detail. In flask culture, when the shaking speed increased from 150 to 220 rpm, the mycelium was altered from a large pellet to a filamentous hypha, and the LME production was almost doubled, while no significant differences were observed among varied filling volumes, which implied a crucial role of shear stress in the morphology and LME production. To confirm this suggestion, experiments with agitation speed ranging from 400 to 1,000 rpm at a fixed aeration rate of 1.0 vvm were conducted in a stirred tank bioreactor. It was found that the morphology became more hairy with reduced pellet size, and the LME production was enhanced threefolds when the agitation speed increased from 400 to 800 rpm. Further experiments by varying initial k L a value at the same agitation speed indicated that oxygen supply only slightly affected the physiological status of ΔstnB1 strain. Altogether, shear stress was identified as a major factor affecting the cell morphology and LME production. The work would be helpful to the production of LME and other secondary metabolites by filamentous microorganism cultivation.
- Published
- 2014
42. Stereospecific Biosynthesis of β-Methyltryptophan from<scp>L</scp>-Tryptophan Features a Stereochemical Switch
- Author
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Qi Fang, Zixin Deng, Dekun Kong, Yi Zou, Haixing Yin, Shuangjun Lin, Linquan Bai, and Zutao Liang
- Subjects
chemistry.chemical_compound ,Stereospecificity ,Biosynthesis ,Chemistry ,Stereochemistry ,Racemases and Epimerases ,Tryptophan ,Stereoisomerism ,General Medicine ,General Chemistry ,Catalysis - Abstract
Make the switch: The three-enzyme cassette MarG/H/I is responsible for stereospecific biosynthesis of β-methyltryptophan from L-tryptophan (1). MarG/I convert 1 into (2S,3R)-β-methyltryptophan, while MarG/I combined with MarH convert 1 into (2S,3S)-β-methyltryptophan. MarH serves as a stereochemical switch by catalyzing the stereoinversion of the β-stereocenter.
- Published
- 2013
43. Characterization of the N-methyltransferase CalM involved in calcimycin biosynthesis by Streptomyces chartreusis NRRL 3882
- Author
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Xiufen Zhou, Jun Yin, Zhijun Wang, Qiulin Wu, Linquan Bai, Derong An, Jingdan Liang, Lixia Gou, Shuangjun Lin, and Zixin Deng
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chemistry.chemical_classification ,Base Sequence ,biology ,Molecular Sequence Data ,Mutagenesis ,Methyltransferases ,General Medicine ,biology.organism_classification ,Biochemistry ,Streptomyces ,Kinetics ,Polyketide ,chemistry.chemical_compound ,Enzyme ,Bacterial Proteins ,Biosynthesis ,chemistry ,Multigene Family ,Gene cluster ,Amino Acid Sequence ,Enzyme kinetics ,Calcimycin - Abstract
Calcimycin is a rare divalent cation specific ionophore antibiotic that has many biochemical and pharmaceutical applications. We have recently cloned and sequenced the Streptomyces chartreusis calcimycin biosynthesis gene cluster as well as identified the genes required for the synthesis of the polyketide backbone of calcimycin. Additional modifying or decorating enzymes are required to convert the polyketide backbone into the biologically active calcimycin. Using targeted mutagenesis of Streptomyces we were able to show that calM from the calcimycin biosynthesis gene cluster is required for calcimycin production. Inactivating calM by PCR targeting, caused high level accumulation of N-demethyl calcimycin. CalM in the presence of S-adenosyl-L-methionine converted N-demethyl calcimycin to calcimycin in vitro. The enzyme was determined to have a kinetic parameter of Km 276 μM, kcat 1.26 min(-1) and kcat/Km 76.2 M(-1) s(-1). These results proved that CalM is a N-methyltransferase that is required for calcimycin biosynthesis, and they set the stage for generating much desired novel calcimycin derivatives by rational genetic and chemical engineering.
- Published
- 2013
44. Biosynthesis and pathway engineering of antifungal polyene macrolides in actinomycetes
- Author
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Eung-Soo Kim, Mi-Jin Lee, Shuangjun Lin, and Dekun Kong
- Subjects
Antifungal ,Antifungal Agents ,Glycosylation ,medicine.drug_class ,Stereochemistry ,Antibiotics ,Bioengineering ,Polyenes ,Hemolysis ,Applied Microbiology and Biotechnology ,Actinobacteria ,chemistry.chemical_compound ,Polyketide ,Cytochrome P-450 Enzyme System ,Biosynthesis ,medicine ,Animals ,biology ,Polyene ,biology.organism_classification ,Anti-Bacterial Agents ,Biosynthetic Pathways ,chemistry ,Biochemistry ,Macrolides ,Genetic Engineering ,Polyketide Synthases ,Biotechnology ,Pathway engineering - Abstract
Polyene macrolides are a large family of natural products typically produced by soil actinomycetes. Polyene macrolides are usually biosynthesized by modular and large type I polyketide synthases (PKSs), followed by several steps of sequential post-PKS modifications such as region-specific oxidations and glycosylations. Although known as powerful antibiotics containing potent antifungal activities (along with additional activities against parasites, enveloped viruses and prion diseases), their high toxicity toward mammalian cells and poor distribution in tissues have led to the continuous identification and structural modification of polyene macrolides to expand their general uses. Advances in in-depth investigations of the biosynthetic mechanism of polyene macrolides and the genetic manipulations of the polyene biosynthetic pathways provide great opportunities to generate new analogues. Recently, a novel class of polyene antibiotics was discovered (a disaccharide-containing NPP) that displays better pharmacological properties such as improved water-solubility and reduced hemolysis. In this review, we summarize the recent advances in the biosynthesis, pathway engineering, and regulation of polyene antibiotics in actinomycetes.
- Published
- 2013
45. Characterization of Streptonigrin Biosynthesis Reveals a Cryptic Carboxyl Methylation and an Unusual Oxidative Cleavage of a N–C Bond
- Author
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Liping Zhang, Xinqiang Xie, Zhang Zhang, Peng Wang, Hairong Cheng, Fei Xu, Zixin Deng, Xinyi He, Meifeng Tao, Shuangjun Lin, Dekun Kong, and Mo Han
- Subjects
Indole test ,Molecular Structure ,Stereochemistry ,Mutant ,General Chemistry ,Methylation ,Protein O-Methyltransferase ,Cleavage (embryo) ,Biochemistry ,Streptomyces ,Catalysis ,chemistry.chemical_compound ,Colloid and Surface Chemistry ,Streptonigrin ,chemistry ,Biosynthesis ,Gene cluster ,Biocatalysis ,Lavendamycin ,Oxidation-Reduction - Abstract
Streptonigrin (STN, 1) is a highly functionalized aminoquinone alkaloid with broad and potent antitumor activity. Here, we reported the biosynthetic gene cluster of STN identified by genome scanning of a STN producer Streptomyces flocculus CGMCC4.1223. This cluster consists of 48 genes determined by a series of gene inactivations. On the basis of the structures of intermediates and shunt products accumulated from five specific gene inactivation mutants and feeding experiments, the biosynthetic pathway was proposed, and the sequence of tailoring steps was preliminarily determined. In this pathway, a cryptic methylation of lavendamycin was genetically and biochemically characterized to be catalyzed by a leucine carboxyl methyltransferase StnF2. A [2Fe-2S](2+) cluster-containing aromatic ring dioxygenase StnB1/B2 system was biochemically characterized to catalyze a regiospecific cleavage of the N-C8' bond of the indole ring of the methyl ester of lavendamycin. This work provides opportunities to illuminate the enzymology of novel reactions involved in this pathway and to create, using genetic and chemo-enzymatic methods, new streptonigrinoid analogues as potential therapeutic agents.
- Published
- 2013
46. The diffusible factor synthase XanB2 is a bifunctional chorismatase that links the shikimate pathway to ubiquinone and xanthomonadins biosynthetic pathways
- Author
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Jianhe Wang, Bangshang Zhu, Changqing Chang, Alan R. Poplawsky, Ji'en Wu, Lian-Hui Zhang, Ya-Wen He, Shuangjun Lin, Jia-Yuan Wang, Tielin Zhou, and Lian Zhou
- Subjects
endocrine system diseases ,biology ,nutritional and metabolic diseases ,Shikimic acid ,biology.organism_classification ,Lyase ,Microbiology ,Xanthomonas campestris ,chemistry.chemical_compound ,chemistry ,Biochemistry ,Biosynthesis ,Xanthomonas ,Gene cluster ,Chorismic acid ,Shikimate pathway ,Molecular Biology - Abstract
The diffusible factor synthase XanB2, originally identified in Xanthomonas campestris pv. campestris (Xcc), is highly conserved across a wide range of bacterial species, but its substrate and catalytic mechanism have not yet been investigated. Here, we show that XanB2 is a unique bifunctional chorismatase that hydrolyses chorismate, the end-product of the shikimate pathway, to produce 3-hydroxybenzoic acid (3-HBA) and 4-HBA. 3-HBA and 4-HBA are respectively associated with the yellow pigment xanthomonadin biosynthesis and antioxidant activity in Xcc. We further demonstrate that XanB2 is a structurally novel enzyme with three putative domains. It catalyses 3-HBA and 4-HBA biosynthesis via a unique mechanism with the C-terminal YjgF-like domain conferring activity for 3-HBA biosynthesis and the N-terminal FGFG motif-containing domain responsible for 4-HBA biosynthesis. Furthermore, we show that Xcc produces coenzyme Q8 (CoQ8) via a new biosynthetic pathway independent of the key chorismate-pyruvate lyase UbiC. XanB2 is the alternative source of 4-HBA for CoQ8 biosynthesis. The similar CoQ8 biosynthetic pathway, xanthomonadin biosynthetic gene cluster and XanB2 homologues are well conserved in the bacterial species within Xanthomonas, Xylella, Xylophilus, Pseudoxanthomonas, Rhodanobacter, Frateuria, Herminiimonas and Variovorax, suggesting that XanB2 may be a conserved metabolic link between the shikimate pathway, ubiquinone and xanthomonadin biosynthetic pathways in diverse bacteria.
- Published
- 2012
47. Enediyne antitumor antibiotic maduropeptin biosynthesis featuring a C-methyltransferase that acts on a CoA-tethered aromatic substrate
- Author
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Jianya Ling, Horsman, Geoffrey P., Sheng-Xiong Huang, Yinggang Luo, Shuangjun Lin, and Ben Shen
- Subjects
Antimitotic agents -- Chemical properties ,Antineoplastic agents -- Chemical properties ,Chromophores -- Structure ,Chromophores -- Chemical properties ,Methylation -- Analysis ,Methyltransferases -- Chemical properties ,Salicylic acid -- Chemical properties ,Chemistry - Abstract
The biosynthetic pathway for the 3,6-dimethylsalicylic acid moiety of the enediyne antitumor antibiotic maduropeptin (MDP) chromophore which is produced by Actinomadura madurae ATCC 39144 is described. The unique property of enzyme MdpB1 to C-methylate a CoA-tethered aromatic acid demonstrated its capability to activating a variety of salicylic acid analogues that could be applied to engineer MDP analogues.
- Published
- 2010
48. Functional Characterization of PyrG, an Unusual Nonribosomal Peptide Synthetase Module from the Pyridomycin Biosynthetic Pathway
- Author
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Shuangjun Lin, Zixin Deng, Lili Li, Tingting Huang, and Nelson L. Brock
- Subjects
0301 basic medicine ,010402 general chemistry ,01 natural sciences ,Biochemistry ,Substrate Specificity ,03 medical and health sciences ,chemistry.chemical_compound ,Biosynthesis ,Bacterial Proteins ,Protein Domains ,Nonribosomal peptide ,Polyketide synthase ,Moiety ,Peptide Synthases ,Molecular Biology ,Adenylylation ,chemistry.chemical_classification ,biology ,INHA ,Organic Chemistry ,Mycobacterium tuberculosis ,0104 chemical sciences ,Amino acid ,Biosynthetic Pathways ,Alcohol Oxidoreductases ,030104 developmental biology ,Enzyme ,chemistry ,biology.protein ,Molecular Medicine ,Oxidoreductases ,Oligopeptides - Abstract
Pyridomycin is an antimycobacterial cyclodepsipeptide assembled by a nonribosomal peptide synthetase/polyketide synthase hybrid system. Analysis of its cluster revealed a nonribosomal peptide synthetase (NRPS) module, PyrG, that contains two tandem adenylation domains and a PKS-type ketoreductase domain. In this study, we biochemically validated that the second A domain recognizes and activates α-keto-β-methylvaleric acid (2-KVC) as the native substrate; the first A domain was not functional but might play a structural role. The KR domain catalyzed the reduction of the 2-KVC tethered to the peptidyl carrier protein of PyrG in the presence of the MbtH family protein, PyrH. PyrG was demonstrated to recognize many amino acids. This substrate promiscuity provides the potential to generate pyridomycin analogues with various enolic acids moiety; this is important for binding InhA, a critical enzyme for cell-wall biosynthesis in Mycobacterium tuberculosis.
- Published
- 2016
49. Identification of (2S,3S)-β-Methyltryptophan as the Real Biosynthetic Intermediate of Antitumor Agent Streptonigrin
- Author
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Shuangjun Lin, Yi Zou, Fei Xu, Liping Zhang, Dekun Kong, Zixin Deng, Zhang Zhang, and Nelson L. Brock
- Subjects
S-Adenosylmethionine ,Transamination ,Mutant ,Antineoplastic Agents ,Stereoisomerism ,Biology ,010402 general chemistry ,01 natural sciences ,Article ,Mass Spectrometry ,chemistry.chemical_compound ,Bacterial Proteins ,Biosynthesis ,Escherichia coli ,Streptonigrin ,Pyridoxal ,Chromatography, High Pressure Liquid ,Antitumor activity ,chemistry.chemical_classification ,Genetics ,Multidisciplinary ,010405 organic chemistry ,Tryptophan ,Methyltransferases ,Streptomyces ,0104 chemical sciences ,Enzyme ,chemistry ,Biochemistry ,Plasmids - Abstract
Streptonigrin is a potent antitumor antibiotic, active against a wide range of mammalian tumor cells. It was reported that its biosynthesis relies on (2S,3R)-β-methyltryptophan as an intermediate. In this study, the biosynthesis of (2S,3R)-β-methyltryptophan and its isomer (2S,3S)-β-methyltryptophan by enzymes from the streptonigrin biosynthetic pathway is demonstrated. StnR is a pyridoxal 5′-phosphate (PLP)-dependent aminotransferase that catalyzes a transamination between L-tryptophan and β-methyl indolepyruvate. StnQ1 is an S-adenosylmethionine (SAM)-dependent C-methyltransferase and catalyzes β-methylation of indolepyruvate to generate (R)-β-methyl indolepyruvate. Although StnR exhibited a significant preference for (S)-β-methyl indolepyruvate over the (R)-epimer, StnQ1 and StnR together catalyze (2S,3R)-β-methyltryptophan formation from L-tryptophan. StnK3 is a cupin superfamily protein responsible for conversion of (R)-β-methyl indolepyruvate to its (S)-epimer and enables (2S,3S)-β-methyltryptophan biosynthesis from L-tryptophan when combined with StnQ1 and StnR. Most importantly, (2S,3S)-β-methyltryptophan was established as the biosynthetic intermediate of the streptonigrin pathway by feeding experiments with a knockout mutant, contradicting the previous proposal that stated (2S,3R)-β-methyltryptophan as the intermediate. These data set the stage for the complete elucidation of the streptonigrin biosynthetic pathway, which would unlock the potential of creating new streptonigrin analogues by genetic manipulation of the biosynthetic machinery.
- Published
- 2016
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50. Xantholipin B produced by the stnR inactivation mutant Streptomyces flocculus CGMCC 4.1223 WJN-1
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Zixin Deng, Sifan Wu, Dan Xie, Tingting Huang, Shuangjun Lin, Xiaozheng Wang, and Jing Wo
- Subjects
Methicillin-Resistant Staphylococcus aureus ,Antifungal Agents ,Antiparasitic ,medicine.drug_class ,Mutant ,Antineoplastic Agents ,Microbial Sensitivity Tests ,Biology ,010402 general chemistry ,Gram-Positive Bacteria ,01 natural sciences ,Streptomyces ,chemistry.chemical_compound ,Structure-Activity Relationship ,Cell Line, Tumor ,Neoplasms ,Drug Discovery ,Xanthone ,medicine ,Structure–activity relationship ,Humans ,Pharmacology ,010405 organic chemistry ,Spectrum Analysis ,Fungi ,biology.organism_classification ,Antimicrobial ,0104 chemical sciences ,Anti-Bacterial Agents ,Biochemistry ,chemistry ,Polyketides ,Antibacterial activity ,Bacteria - Abstract
Xantholipin is a polycyclic xanthone antibiotic that exhibits potent cytotoxic and antibacterial activity. In this study, a new xanthone-type antibiotic, xantholipin B (1), was isolated for the first time along with its known derivative, xantholipin (2), from strain WJN-1, an aminotransferase inactivation mutant of the streptonigrin-producer Streptomyces flocculus CGMCC 4.1223. The structure of 1 was established based on spectroscopic analysis and supports the previously proposed biosynthetic pathway as a key intermediate of 2. Moreover, 1 showed 3- to 10-fold greater cytotoxicity than 2 against a select panel of human cancer cell lines. In addition, 1 demonstrated powerful antimicrobial activity against both Gram-positive bacteria and fungi. Importantly, both 1 and 2 inhibited the methicillin-resistant strain Staphylococcus aureus Mu50, with the MIC value of 0.025 μg ml-1. The new structural features of 1 enrich the structural diversity of xantholipin family compounds and shed new light on the structure-activity relationship of 1 as a promising antitumor drug candidate.
- Published
- 2016
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