Production of chain-extended cinnamoyl compounds by overexpressing two adjacent cluster-situated LuxR regulators in Streptomyces globisporus C-1027

Natural products from microorganisms are important sources for drug discovery. With the development of high-throughput sequencing technology and bioinformatics, a large amount of uncharacterized biosynthetic gene clusters (BGCs) in microorganisms have been found, which show the potential for novel natural product production. Nine BGCs containing PKS and/or NRPS in Streptomyces globisporus C-1027 were transcriptionally low/silent under the experimental fermentation conditions, and the products of these clusters are unknown. Thus, we tried to activate these BGCs to explore cryptic products of this strain. We constructed the cluster-situated regulator overexpressing strains which contained regulator gene(s) under the control of the constitutive promoter ermE*p in S. globisporus C-1027. Overexpression of regulators in cluster 26 resulted in significant transcriptional upregulation of biosynthetic genes. With the separation and identification of products from the overexpressing strain OELuxR1R2, three ortho-methyl phenyl alkenoic acids (compounds 1-3) were obtained. Gene disruption showed that compounds 1 and 2 were completely abolished in the mutant GlaEKO, but were hardly affected by deletion of the genes orf3 or echA in cluster 26. The type II PKS biosynthetic pathway of chain-extended cinnamoyl compounds was deduced by bioinformatics analysis. This study showed that overexpression of the two adjacent cluster-situated LuxR regulator(s) is an effective strategy to connect the orphan BGC to its products.

Automated summary

Natural products from microorganisms play a significant role in drug discovery. This study focuses on uncharacterized biosynthetic gene clusters in Streptomyces globisporus C-1027, and attempts to activate these clusters to explore unknown products. By overexpressing regulators in cluster 26, the transcriptional upregulation of biosynthetic genes was achieved, leading to the identification of three ortho-methyl phenyl alkenoic acids. This study demonstrates that overexpression of cluster-situated regulators is an effective strategy to connect orphan biosynthetic gene clusters with their products.