Cloning and Heterologous Expression of the Macrotetrolide Biosynthetic Gene Cluster Revealed a Novel Polyketide Synthase that Lacks an Acyl Carrier Protein

2001 ◽  
Vol 123 (14) ◽  
pp. 3385-3386 ◽  
Author(s):  
Hyung-Jin Kwon ◽  
Wyatt C. Smith ◽  
Longkuan Xiang ◽  
Ben Shen
Keyword(s):  
Heterologous Expression ◽  
Gene Cluster ◽  
Polyketide Synthase ◽  
Acyl Carrier Protein ◽  
Biosynthetic Gene Cluster ◽  
Carrier Protein ◽  
Biosynthetic Gene

Cloning, sequencing and heterologous expression of the medermycin biosynthetic gene cluster of Streptomyces sp. AM-7161: towards comparative analysis of the benzoisochromanequinone gene clusters

Microbiology ◽  
2003 ◽  
Vol 149 (7) ◽  
pp. 1633-1645 ◽  
Author(s):  
Koji Ichinose ◽  
Makoto Ozawa ◽  
Keiko Itou ◽  
Kanako Kunieda ◽  
Yutaka Ebizuka
Keyword(s):  
Heterologous Expression ◽  
Gene Cluster ◽  
Polyketide Synthase ◽  
Acyl Carrier Protein ◽  
Streptomyces Coelicolor ◽  
Gene Clusters ◽  
Biosynthetic Gene Cluster ◽  
Carrier Protein ◽  
Biosynthetic Gene ◽  
Six Genes

Medermycin is a Streptomyces aromatic C-glycoside antibiotic classified in the benzoisochromanequinones (BIQs), which presents several interesting biosynthetic problems concerning polyketide synthase (PKS), post-PKS tailoring and deoxysugar pathways. The biosynthetic gene cluster for medermycin (the med cluster) was cloned from Streptomyces sp. AM-7161. Completeness of the clone was proved by the heterologous expression of a cosmid carrying the entire med cluster in Streptomyces coelicolor CH999 to produce medermycin. The DNA sequence of the cosmid (36 202 bp) revealed 34 complete ORFs, with an incomplete ORF at either end. Functional assignment of the deduced products was made for PKS and biosynthetically related enzymes, tailoring steps including strereochemical control, oxidation, angolosamine pathway, C-glycosylation, and regulation. The med cluster was estimated to be about 30 kb long, covering 29 ORFs. An unusual characteristic of the cluster is the disconnected organization of the minimal PKS genes: med-ORF23 encoding the acyl carrier protein is 20 kb apart from med-ORF1 and med-ORF2 for the two ketosynthase components. Secondly, the six genes (med-ORF14, 15, 16, 17, 18 and 20) for the biosynthesis of the deoxysugar, angolosamine, are all contiguous. Finally, the finding of a glycosyltransferase gene, med-ORF8, suggests a possible involvement of conventional C-glycosylation in medermycin biosynthesis. Comparison among the three complete BIQ gene clusters – med and those for actinorhodin (act) and granaticin (gra) – revealed some common genes whose deduced functions are unavailable from database searches (the ‘unknowns’). An example is med-ORF5, a homologue of actVI-ORF3 and gra-ORF18, which was highlighted by a recent proteomic analysis of S. coelicolor A3(2).

scholarly journals Utilization of the Methoxymalonyl-Acyl Carrier Protein Biosynthesis Locus for Cloning the Oxazolomycin Biosynthetic Gene Cluster from Streptomyces albus JA3453

2006 ◽  
Vol 188 (11) ◽  
pp. 4142-4147 ◽  
Author(s):  
Chunhua Zhao ◽  
Jianhua Ju ◽  
Steven D. Christenson ◽  
Wyatt C. Smith ◽  
Danfeng Song ◽  
...  
Keyword(s):  
Gene Cluster ◽  
Acyl Carrier Protein ◽  
Protein Biosynthesis ◽  
Biosynthetic Gene Cluster ◽  
Carrier Protein ◽  
Biosynthetic Gene ◽  
Degenerate Primers ◽  
Streptomyces Albus ◽  
Genes Encoding ◽  
Antiviral Activities

ABSTRACT Oxazolomycin (OZM), a hybrid peptide-polyketide antibiotic, exhibits potent antitumor and antiviral activities. Using degenerate primers to clone genes encoding methoxymalonyl-acyl carrier protein (ACP) biosynthesis as probes, a 135-kb DNA region from Streptomyces albus JA3453 was cloned and found to cover the entire OZM biosynthetic gene cluster. The involvement of the cloned genes in OZM biosynthesis was confirmed by deletion of a 12-kb DNA fragment containing six genes for methoxymalonyl-ACP biosynthesis from the specific region of the chromosome, as well as deletion of the ozmC gene within this region, to generate OZM-nonproducing mutants.

scholarly journals Utilization of the Methoxymalonyl-Acyl Carrier Protein Biosynthesis Locus for Cloning of the Tautomycin Biosynthetic Gene Cluster from Streptomyces spiroverticillatus

2006 ◽  
Vol 188 (11) ◽  
pp. 4148-4152 ◽  
Author(s):  
Wenli Li ◽  
Jianhua Ju ◽  
Hiroyuki Osada ◽  
Ben Shen
Keyword(s):  
Gene Cluster ◽  
Protein Phosphatase ◽  
Acyl Carrier Protein ◽  
Protein Biosynthesis ◽  
Acyl Chain ◽  
Biosynthetic Gene Cluster ◽  
Carrier Protein ◽  
Biosynthetic Gene ◽  
Degenerate Primers ◽  
Polyketide Chain

ABSTRACT Tautomycin (TTM), a potent protein phosphatase inhibitor, consists of a polyketide chain containing a spiroketal moiety and an acyl chain bearing a dialkylmaleic anhydride structure. PCR using degenerate primers was used to clone genes from Streptomyces spiroverticillatus for formation of the methoxymalonyl-acyl carrier protein. This locus was found to contain five genes (ttmC, ttmA, ttmD, ttmB, and ttmE), one of which was used as a probe to clone the 110-kb TTM biosynthetic gene cluster. The involvement of the ttmA gene in TTM biosynthesis was confirmed by gene inactivation and mutation complementation experiments.

scholarly journals Identification and Heterologous Expression of the Kendomycin B Biosynthetic Gene Cluster from Verrucosispora sp. SCSIO 07399

Marine Drugs ◽  
2021 ◽  
Vol 19 (12) ◽  
pp. 673
Author(s):  
Jiang Chen ◽  
Shanwen Zhang ◽  
Yingying Chen ◽  
Xinpeng Tian ◽  
Yucheng Gu ◽  
...  
Keyword(s):  
Heterologous Expression ◽  
Gene Cluster ◽  
Polyketide Synthase ◽  
Regulatory Gene ◽  
Biosynthetic Gene Cluster ◽  
Chain Elongation ◽  
Type I ◽  
Biosynthetic Gene ◽  
Type Iii Polyketide Synthase ◽  
Type Iii

Verrucosispora sp. SCSIO 07399, a rare marine-derived actinomycete, produces a set of ansamycin-like polyketides kendomycin B–D (1–3) which possess potent antibacterial activities and moderate tumor cytotoxicity. Structurally, kendomycin B–D contain a unique aliphatic macrocyclic ansa scaffold in which the highly substituted pyran ring is connected to the quinone moiety. In this work, a type I/type III polyketide synthase (PKS) hybrid biosynthetic gene cluster coding for assembly of kendomycin B (kmy), and covering 33 open reading frames, was identified from Verrucosispora sp. SCSIO 07399. The kmy cluster was found to be essential for kendomycin B biosynthesis as verified by gene disruption and heterologous expression. Correspondingly, a biosynthetic pathway was proposed based on bioinformatics, cluster alignments, and previous research. Additionally, the role of type III PKS for generating the precursor unit 3,5-dihydroxybenzoic acid (3,5-DHBA) was demonstrated by chemical complementation, and type I PKS executed the polyketide chain elongation. The kmy cluster was found to contain a positive regulatory gene kmy4 whose regulatory effect was identified using real-time quantitative PCR (RT-qPCR). These advances shed important new insights into kendomycin B biosynthesis and help to set the foundation for further research aimed at understanding and exploiting the carbacylic ansa scaffold.

Direct Cloning, Genetic Engineering, and Heterologous Expression of the Syringolin Biosynthetic Gene Cluster inE. colithrough Red/ET Recombineering

ChemBioChem ◽  
2012 ◽  
Vol 13 (13) ◽  
pp. 1946-1952 ◽  
Author(s):  
Xiaoying Bian ◽  
Fan Huang ◽  
Francis A. Stewart ◽  
Liqiu Xia ◽  
Youming Zhang ◽  
...  
Keyword(s):  
Genetic Engineering ◽  
Heterologous Expression ◽  
Gene Cluster ◽  
Biosynthetic Gene Cluster ◽  
Biosynthetic Gene ◽  
Direct Cloning

Identification of the kinanthraquinone biosynthetic gene cluster by expression of an atypical response regulator

2021 ◽  
Vol 85 (3) ◽  
pp. 714-721
Author(s):  
Risa Takao ◽  
Katsuyuki Sakai ◽  
Hiroyuki Koshino ◽  
Hiroyuki Osada ◽  
Shunji Takahashi
Keyword(s):  
Heterologous Expression ◽  
Gene Cluster ◽  
Secondary Metabolite ◽  
Response Regulator ◽  
Bac Library ◽  
Gene Clusters ◽  
Biosynthetic Gene Cluster ◽  
Gene Organization ◽  
Biosynthetic Gene ◽  
Streptomyces Sp

ABSTRACT Recent advances in genome sequencing have revealed a variety of secondary metabolite biosynthetic gene clusters in actinomycetes. Understanding the biosynthetic mechanism controlling secondary metabolite production is important for utilizing these gene clusters. In this study, we focused on the kinanthraquinone biosynthetic gene cluster, which has not been identified yet in Streptomyces sp. SN-593. Based on chemical structure, 5 type II polyketide synthase gene clusters were listed from the genome sequence of Streptomyces sp. SN-593. Among them, a candidate gene cluster was selected by comparing the gene organization with grincamycin, which is synthesized through an intermediate similar to kinanthraquinone. We initially utilized a BAC library for subcloning the kiq gene cluster, performed heterologous expression in Streptomyces lividans TK23, and identified the production of kinanthraquinone and kinanthraquinone B. We also found that heterologous expression of kiqA, which belongs to the DNA-binding response regulator OmpR family, dramatically enhanced the production of kinanthraquinones.

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