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The cinnamoyl lipid compound youssoufene A1 (1), featuring a unique dearomatic carbon‐bridged dimeric skeleton, exhibits increased inhibition against multidrug resistant Enterococcus faecalis as compared to monomeric youssoufenes. However, the formation process of this intriguing dearomatization/dimerization remains unknown. In this study, an unusual "gene‐within‐gene" thioesterase (TE) gene ysfF was functionally characterized. The gene was found to naturally encodes two proteins, an entire YsfF with α/β‐hydrolase and 4‐hydroxybenzoyl‐CoA thioesterase (4‐HBT)‐like enzyme domains, and a nested YsfFHBT (4‐HBT‐like enzyme). Using an intracellular tagged carrier‐protein tracking (ITCT) strategy, in vitro reconstitution and in vivo experiments, we found that: i) both domains of YsfF displayed thioesterase activities; ii) YsfF/YsfFHBT could accomplish the 6π‐electrocyclic ring closure for benzene ring formation; and iii) YsfF and cyclase YsfX together were responsible for the ACP‐tethered dearomatization/dimerization process, possibly through an unprecedented Michael‐type addition reaction. Moreover, site‐directed mutagenesis experiments demonstrated that N301, E483 and H566 of YsfF are critical residues for both the 6π‐electrocyclization and dimerization processes. This study enhances our understanding of the multifunctionality of the TE protein family. [ABSTRACT FROM AUTHOR]

As a unique structural moiety in natural products, cinnamoyl lipids (CLs), are proposed to be assembled by unusual type II polyketide synthases (PKSs). Herein, we demonstrate that the assembly of the CL compounds youssoufenes is accomplished by a PKS system that uniquely harbors three phylogenetically different ketosynthase/chain length factor (KS/CLF) complexes (YsfB/C, YsfD/E, and YsfJ/K). Through in vivo gene inactivation and in vitro reconstitution, as well as an intracellular tagged carrier‐protein tracking (ITCT) strategy developed in this study, we successfully elucidated the isomerase‐dependent ACP‐tethered polyunsaturated chain elongation process. The three KS/CLFs were revealed to modularly assemble different parts of the youssoufene skeleton, during which benzene ring closure happens right after the formation of an ACP‐tethered C18 polyene. Of note, the ITCT strategy could significantly contribute to the elucidation of other carrier‐protein‐dependent biosynthetic machineries. [ABSTRACT FROM AUTHOR]

Nonsense-mediated decay (NMD) mechanism, also called mRNA surveillance, is a universal mRNA degradation pathway in eukaryotes. Hundreds of genes can be regulated by NMD whether in single-celled or higher organisms. There have been many studies on NMD and NMD factors (Upf proteins) with regard to their crucial roles in mRNA decay, especially in mammals and yeast. However, research focusing on NMD in plant is still lacking compared to the research that has been dedicated to NMD in mammals and yeast. Even so, recent study has shown that NMD factors in Arabidopsis can provide resistance against biotic and abiotic stresses. This discovery and its associated developments have given plant NMD mechanism a new outlook and since then, more and more research has focused on this area. In this review, we focused mainly on the distinctive NMD micromechanism and functions of Upf proteins in plant with references to the role of mRNA surveillance in mammals and yeast. We also highlighted recent insights into the roles of premature termination codon location, trans-elements and functions of other NMD factors to emphasize the particularity of plant NMD. Furthermore, we also discussed conventional approaches and neoteric methods used in plant NMD researches. [ABSTRACT FROM AUTHOR]

The gene, sabR, encoding a receptor for γ-butyrolactone, was cloned from the genomic DNA of Streptomyces ansochromogenes 7100. Its deduced protein shows strong homology to several γ-butyrolactone-binding proteins in Streptomyces. Disruption of sabR retarded nikkomycin production in liquid media containing glucose or glycerol as carbon source. Sporulation of sabR disruption mutants was earlier than the parent strain on solid media with glucose or glycerol as carbon source. However, disruption of sabR had no effect on either nikkomycin production or sporulation on media containing mannitol as carbon source, suggesting that sabR is a pleiotropic regulatory gene that controls the onset of nikkomycin production and sporulation in S. ansochromogenes and is related to the utilization of carbon source. [ABSTRACT FROM AUTHOR]

Focuses on a study which examined the sequencing and function of the sanB gene, which is involved in the nikkomycin biosynthesis of Streptomyces ansochromogenes. Materials and methods; Results; Discussion.

Background: Polyene antibiotics are important as antifungal medicines albeit with serious side effects such as nephrotoxicity. Reedsmycin (RDM) A (1), produced by marine-derived Streptomyces youssoufiensis OUC6819, is a non-glycosylated polyene macrolide antibiotic with antifungal activity comparable to that of clinically used nystatin. To elucidate its biosynthetic machinery, herein, the rdm biosynthetic gene cluster was cloned and characterized. Results: The rdm cluster is located within a 104 kb DNA region harboring 21 open reading frames (ORFs), among which 15 ORFs were designated as rdm genes. The assembly line for RDM A is proposed on the basis of module and domain analysis of the polyketide synthetases (PKSs) RdmGHIJ, which catalyze 16 rounds of decarboxylative condensation using malonyl-CoA as the starter unit (loading module), two methylmalonyl-CoA (module 1 and 2), and fourteen malonyl-CoA (module 3–16) as extender units successively. However, the predicted substrate specificity of AT0 in the loading module is methylmalonyl-CoA instead of malonyl-CoA. Interestingly, the rdm cluster contains a five-gene regulation system RdmACDEF, which is different from other reported polyene gene clusters. In vivo experiments demonstrated the XRE family regulator RdmA and the PAS/LuxR family regulator RdmF function in negative and positive manner, respectively. Notably, inactivation of rdmA and overexpression of rdmF led to increased production of RDM A by ~ 2.0-fold and ~ 2.5-fold, reaching yields of 155.3 ± 1.89 and 184.8 ± 9.93 mg/L, respectively. Conclusions: Biosynthesis of RDM A is accomplished on a linear assembly line catalyzed by Rdm PKSs harboring a unique AT0 under the control of a complex regulatory system. These findings enable generation of new biologically active RDM derivatives at high yield and with improved properties by engineered biosynthesis. [ABSTRACT FROM AUTHOR]

Drimentines (DMTs) are a family of bacterial alkaloids featuring a 2,5-diketopiperazine (2, 5-DKP) ring with diverse bioactivities. In this study, we characterized the biosynthetic pathway of DMT B through gene combinations and biochemical investigations. While cyclo (L -Trp- L -Pro) (cWP) is synthesized by a tRNA-dependent cyclodipeptide synthase (CDPS) DmtB2, further tailoring steps are achieved sequentially through cyclodipeptide oxidases DmtD2_E2, prenyltransferase DmtC2, and terpene cyclase DmtA2. Our results deepen the understanding of the biosynthetic logic of DMT compounds, providing additional biological information for the structural diversification of terpenylated DKP compounds. [Display omitted] [ABSTRACT FROM AUTHOR]