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

Jiang Chen,Shanwen Zhang,Jianhua Ju,Yingying Chen,Yucheng Gu,Xinpeng Tian

doi:10.3390/md19120673

Abstract

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.

Highlights

The kendomycins are ansamycin-type polyketides possessing an all-carbon macrocyclic ansa skeleton characterized by a highly substituted pyran ring and a quinone methide chromophore
The main difference between the kmy and ken clusters is that the kmy cluster lacks a methyltransferase gene thought to provide a para-methyl moiety of the starter unit; this distinction is reflected in the solved structure of kendomycin B (Figure S1)
A type I/type III polyketide synthase (PKS) hybrid biosynthetic gene cluster encoding the assembly of kendomycin B was identified from marine-derived Verrucosispora sp

Summary

Introduction

The kendomycins are ansamycin-type polyketides possessing an all-carbon macrocyclic ansa skeleton characterized by a highly substituted pyran ring and a quinone methide chromophore In addition to their complex and unique structures, members of the compound class display remarkable antibacterial, anti-osteoporotic, and anticancer activities; they show great promise as the drug leads for a wide array of clinical applications [1]. By virtue of these properties, the kendomycin class of natural products has long held the interest of the medicinal and synthetic organic communities. Biologists have long pondered the biosynthetic chronology and enzymology of the post-modifications, underlying the intricate cyclization reactions necessary to attain the complete natural product

Methods

Results

Conclusion