7 results on '"Ding, Junmei"'
Search Results
2. Characterization of a Glycoside Hydrolase Family 27 α-Galactosidase from Pontibacter Reveals Its Novel Salt–Protease Tolerance and Transglycosylation Activity
- Author
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Chunyan Li, Xu Bo, Ding Junmei, Junpei Zhou, Junjun Li, Xianghua Tang, Yu Liu, Rui Zhang, Qian Lu, Qian Wu, Zunxi Huang, and Han Nanyu
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0301 basic medicine ,Sucrose ,Glycosylation ,medicine.medical_treatment ,Gene Expression ,Sodium Chloride ,03 medical and health sciences ,chemistry.chemical_compound ,Pentanols ,Raffinose ,Escherichia coli ,medicine ,Trypsin ,Glycoside hydrolase ,Cloning, Molecular ,chemistry.chemical_classification ,Guar gum ,Protease ,Alpha-galactosidase ,biology ,Bacteroidetes ,General Chemistry ,Recombinant Proteins ,030104 developmental biology ,Enzyme ,chemistry ,Biochemistry ,alpha-Galactosidase ,biology.protein ,Locust bean gum ,General Agricultural and Biological Sciences ,medicine.drug - Abstract
α-Galactosidases are of great interest in various applications. A glycoside hydrolase family 27 α-galactosidase was cloned from Pontibacter sp. harbored in a saline soil and expressed in Escherichia coli. The purified recombinant enzyme (rAgaAHJ8) was little or not affected by 3.5-30.0% (w/v) NaCl, 10.0-100.0 mM Pb(CH3COO)2, 10.0-60.0 mM ZnSO4, or 8.3-100.0 mg mL(-1) trypsin and by most metal ions and chemical reagents at 1.0 and 10.0 mM concentrations. The degree of synergy on enzymatic degradation of locust bean gum and guar gum by an endomannanase and rAgaAHJ8 was 1.22-1.54. In the presence of trypsin, the amount of reducing sugars released from soybean milk treated by rAgaAHJ8 was approximately 3.8-fold compared with that treated by a commercial α-galactosidase. rAgaAHJ8 showed transglycosylation activity when using sucrose, raffinose, and 3-methyl-1-butanol as the acceptors. Furthermore, potential factors for salt adaptation of the enzyme were presumed.
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- 2016
3. Distinctive molecular and biochemical characteristics of a glycoside hydrolase family 20 β-N-acetylglucosaminidase and salt tolerance
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Zunxi Huang, Zhou Junpei, Bo Xu, Rui Liu, Ding Junmei, Qian Wu, Zhifeng Song, Nanyu Han, Rui Zhang, Junjun Li, and Tang Xianghua
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0301 basic medicine ,Glycoside Hydrolases ,Protein Conformation ,lcsh:Biotechnology ,Microbacterium ,Lysine ,Chitobiose ,Acetylglucosamine ,Substrate Specificity ,Microbiology ,03 medical and health sciences ,chemistry.chemical_compound ,Chitin ,lcsh:TP248.13-248.65 ,Acetylglucosaminidase ,Enzyme Stability ,Aspartic acid ,Glycoside hydrolase ,chemistry.chemical_classification ,Binding Sites ,biology ,Salt Tolerance ,biology.organism_classification ,Enzyme assay ,Activity ,Actinobacteria ,Enzyme Activation ,N-Acetylglucosaminide tolerance ,030104 developmental biology ,Enzyme ,Biochemistry ,chemistry ,biology.protein ,Salts ,β-N-Acetylglucosaminidase ,Protein Binding ,Research Article ,Biotechnology - Abstract
Background Enzymatic degradation of chitin has attracted substantial attention because chitin is an abundant renewable natural resource, second only to lignocellulose, and because of the promising applications of N-acetylglucosamine in the bioethanol, food and pharmaceutical industries. However, the low activity and poor tolerance to salts and N-acetylglucosamine of most reported β-N-acetylglucosaminidases limit their applications. Mining for novel enzymes from new microorganisms is one way to address this problem. Results A glycoside hydrolase family 20 (GH 20) β-N-acetylglucosaminidase (GlcNAcase) was identified from Microbacterium sp. HJ5 harboured in the saline soil of an abandoned salt mine and was expressed in Escherichia coli. The purified recombinant enzyme showed specific activities of 1773.1 ± 1.1 and 481.4 ± 2.3 μmol min−1 mg−1 towards p-nitrophenyl β-N-acetylglucosaminide and N,N'-diacetyl chitobiose, respectively, a V max of 3097 ± 124 μmol min−1 mg−1 towards p-nitrophenyl β-N-acetylglucosaminide and a K i of 14.59 mM for N-acetylglucosamine inhibition. Most metal ions and chemical reagents at final concentrations of 1.0 and 10.0 mM or 0.5 and 1.0% (v/v) had little or no effect (retaining 84.5 − 131.5% activity) on the enzyme activity. The enzyme can retain more than 53.6% activity and good stability in 3.0–20.0% (w/v) NaCl. Compared with most GlcNAcases, the activity of the enzyme is considerably higher and the tolerance to salts and N-acetylglucosamine is much better. Furthermore, the enzyme had higher proportions of aspartic acid, glutamic acid, alanine, glycine, random coils and negatively charged surfaces but lower proportions of cysteine, lysine, α-helices and positively charged surfaces than its homologs. These molecular characteristics were hypothesised as potential factors in the adaptation for salt tolerance and high activity of the GH 20 GlcNAcase. Conclusions Biochemical characterization revealed that the GlcNAcase had novel salt–GlcNAc tolerance and high activity. These characteristics suggest that the enzyme has versatile potential in biotechnological applications, such as bioconversion of chitin waste and the processing of marine materials and saline foods. Molecular characterization provided an understanding of the molecular–function relationships for the salt tolerance and high activity of the GH 20 GlcNAcase. Electronic supplementary material The online version of this article (doi:10.1186/s12896-017-0358-1) contains supplementary material, which is available to authorized users.
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- 2017
4. Genetic diversity of catechol 1,2-dioxygenase in the fecal microbial metagenome
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Ding Junmei, Caiyun Xiong, Meng Deng, Nanyu Han, Tang Xianghua, Junjun Li, Junpei Zhou, Yunjuan Yang, Qian Wu, Zunxi Huang, and Bo Xu
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0301 basic medicine ,Microbial DNA ,Catechols ,medicine.disease_cause ,Applied Microbiology and Biotechnology ,Polymerase Chain Reaction ,Microbiology ,03 medical and health sciences ,chemistry.chemical_compound ,Feces ,medicine ,Escherichia coli ,Catechol 1,2-dioxygenase ,Animals ,Enzyme kinetics ,Cloning, Molecular ,Phylogeny ,DNA Primers ,chemistry.chemical_classification ,Catechol ,Acinetobacter ,Genetic Variation ,General Medicine ,Catechol 1,2-Dioxygenase ,Lorisidae ,Kinetics ,030104 developmental biology ,Enzyme ,Biochemistry ,chemistry ,Metagenomics ,Metagenome ,Heterologous expression - Abstract
Catechol 1,2-dioxygenase is the key enzyme that catalyzes the cleavage of the aromatic ring of catechol. We explored the genetic diversity of catechol 1,2-dioxygenase in the fecal microbial metagenome by PCR with degenerate primers. A total of 35 gene fragments of C12O were retrieved from microbial DNA in the feces of pygmy loris. Based on phylogenetic analysis, most sequences were closely related to C12O sequences from Acinetobacter. A full-length C12O gene was directly cloned, heterologously expressed in Escherichia coli, and biochemically characterized. Purified catPL12 had optimum pH and temperature pH 8.0 and 25 °C and retained 31 and 50% of its maximum activity when assayed at 0 and 35 °C, respectively. The enzyme was stable at 25 and 37 °C, retaining 100% activity after pre-incubation for 1 h. The kinetic parameters of catPL12 were determined. The enzyme had apparent Km of 67 µM, Vmax of 7.3 U/mg, and kcat of 4.2 s-1 for catechol, and the cleavage activities for 3-methylcatechol, 4-methylcatechol, and 4-chlorocatechol were much less than for catechol, and no activity with hydroquinone or protocatechuate was detected. This study is the first to report the molecular and biochemical characterizations of a cold-adapted catechol 1,2-dioxygenase from a fecal microbial metagenome.
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- 2017
5. Characterization of a novel low-temperature-active, alkaline and sucrose-tolerant invertase
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Han Nanyu, Tang Xianghua, Yajie Gao, He Limei, Huang Zunxi, Xu Bo, Ding Junmei, Qian Wu, Rui Zhang, Junpei Zhou, and Junjun Li
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0106 biological sciences ,0301 basic medicine ,Sucrose ,Glycosylation ,Protein Conformation ,Glycine ,Bacillus ,01 natural sciences ,Article ,Substrate Specificity ,03 medical and health sciences ,chemistry.chemical_compound ,Hydrolysis ,Sequence Analysis, Protein ,Catalytic Domain ,Escherichia coli ,Glycoside hydrolase ,Enzyme kinetics ,Proline ,chemistry.chemical_classification ,Multidisciplinary ,beta-Fructofuranosidase ,Temperature ,Hydrogen-Ion Concentration ,Recombinant Proteins ,Kinetics ,030104 developmental biology ,Enzyme ,Invertase ,chemistry ,Biochemistry ,Thermodynamics ,010606 plant biology & botany - Abstract
A glycoside hydrolase family 32 invertase from Bacillus sp. HJ14 was expressed in Escherichia coli. The purified recombinant enzyme (rInvHJ14) showed typical biochemical properties of low-temperature-active and alkaline enzymes: (i) rInvHJ14 was active and stable in the range of pH 7.0–9.5 with an apparent pH optimum of 8.0; (ii) rInvHJ14 was most active but not stable at 30–32.5 °C, with 19.7, 48.2 and 82.1% of its maximum activity when assayed at 0, 10 and 20 °C, respectively and the Ea, ΔG* (30 °C), Km (30 °C) and kcat (30 °C) values for hydrolysis of sucrose by rInvHJ14 was 47.6 kJ mol−1, 57.6 kJ mol−1, 62.9 mM and 746.2 s−1, respectively. The enzyme also showed strong sucrose tolerance. rInvHJ14 preserved approximately 50% of its highest activity in the presence of 2045.0 mM sucrose. Furthermore, potential factors for low-temperature-active and alkaline adaptations of rInvHJ14 were presumed. Compared with more thermostable homologs, rInvHJ14 has a higher frequency of glycine residues and a longer loop but a lower frequency of proline residues (especially in a loop) in the catalytic domain. The catalytic pockets of acid invertases were almost negatively charged while that of alkaline rInvHJ14 was mostly positively charged.
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- 2016
6. NaCl-, protease-tolerant and cold-active endoglucanase from Paenibacillus sp. YD236 isolated from the feces of Bos frontalis
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Junjun Li, Junpei Zhou, Mingjie Dong, Huang Zunxi, Tang Xianghua, Yunjuan Yang, Shen Jidong, Wu Qian, Han Nanyu, Xu Bo, Ding Junmei, and Yuelin Mu
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0301 basic medicine ,Bos frontalis ,Starch ,medicine.medical_treatment ,Cellulase ,Microbiology ,03 medical and health sciences ,Paenibacillus ,chemistry.chemical_compound ,Laminarin ,medicine ,Endoglucanase ,Glucan ,chemistry.chemical_classification ,Multidisciplinary ,Protease ,biology ,Research ,Enzyme characterization ,biology.organism_classification ,Trypsin ,Enzyme assay ,030104 developmental biology ,chemistry ,biology.protein ,Heterologous expression ,medicine.drug - Abstract
Bos frontalis, which consumes bamboo and weeds, may have evolved unique gastrointestinal microorganisms that digest cellulase. A Paenibacillus sp. YD236 strain was isolated from B. frontalis feces, from which a GH8 endoglucanase gene, pglue8 (1107 bp, 54.5 % GC content), encoding a 368-residue polypeptide (PgluE8, 40.4 kDa) was cloned. PgluE8 efficiently hydrolyzed barley-β-d-glucan followed by CMC-Na, soluble starch, laminarin, and glucan from black yeast optimally at pH 5.5 and 50 °C, and retained 78.6, 41.6, and 34.5 % maximum activity when assayed at 20, 10, and 0 °C, respectively. Enzyme activity remained above 176.6 % after treatment with 10.0 mM β-mercaptoethanol, and was 83.0, 78, and 56 % after pre-incubation in 30 % (w/v) NaCl, 16.67 mg/mL trypsin, and 160.0 mg/mL protease K, respectively. Cys23 and Cys364 residues were critical for PgluE8 activity. pglue8, identified from B. frontalis feces for the first time in this study, is a potential alternative for applications including food processing, washing, and animal feed preparation.
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- 2016
7. Characterization of two glycoside hydrolase family 36 α-galactosidases: novel transglycosylation activity, lead-zinc tolerance, alkaline and multiple pH optima, and low-temperature activity
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Yiyan Wang, Junpei Zhou, Tang Xianghua, Rui Zhang, Junjun Li, Zunxi Huang, Xu Bo, Ding Junmei, Qian Wu, and Qian Lu
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0301 basic medicine ,Stereochemistry ,medicine.disease_cause ,Streptomyces ,Analytical Chemistry ,03 medical and health sciences ,chemistry.chemical_compound ,Hydrolysis ,medicine ,Glycoside hydrolase ,Enzyme kinetics ,Glycosides ,Escherichia coli ,chemistry.chemical_classification ,Alpha-galactosidase ,biology ,Temperature ,Glycoside ,Galactose ,General Medicine ,biology.organism_classification ,Zinc ,030104 developmental biology ,Biochemistry ,chemistry ,Lead ,alpha-Galactosidase ,biology.protein ,Food Science - Abstract
Two α-galactosidases, AgaAJB07 from Mesorhizobium and AgaAHJG4 from Streptomyces, were expressed in Escherichia coli. Recombinant AgaAJB07 showed a 2.9-fold and 22.6-fold increase in kcat with a concomitant increase of 2.3-fold and 16.3-fold in Km in the presence of 0.5mM ZnSO4 and 30.0mM Pb(CH3COO)2, respectively. Recombinant AgaAHJG4 showed apparent optimal activity at pH 8.0 in McIlvaine or Tris-HCl buffer and 9.5 in glycine-NaOH or HCl-borax-NaOH buffer, retention of 23.6% and 43.2% activity when assayed at 10 and 20°C, respectively, and a half-life of approximately 2min at 50°C. The activation energies for p-nitrophenyl-α-d-galactopyranoside hydrolysis by AgaAJB07 and AgaAHJG4 were 71.9±0.8 and 48.2±2.0kJmol(-1), respectively. Both AgaAJB07 and AgaAHJG4 exhibited transglycosylation activity, but they required different acceptors and produced different compounds. Furthermore, potential factors for alkaline and multiple pH optima and low-temperature adaptations of AgaAHJG4 were presumed.
- Published
- 2015
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