Investigation of the biosynthetic gene cluster of trialkyl-substituted aromatic polyketide gombapyrones from Streptomyces rubellomurinus

Abstract:

[Objective] To identify the biosynthetic gene cluster (BGC) of gombapyrones (GOMs), the trialkyl-substituted aromatic polyketides derived from Streptomyces rubellomurinus ATCC 31215, and deduce the biosynthetic pathway. [Methods] GOM-B and GOM-D were extracted for the large-scale fermentation broth of S. rubellomurinus ATCC 31215. The P450 monooxygenase catalyzing the polyene chain aromatization is conserved in the biosynthetic pathways of trialkyl-substituted aromatic polyketides in bacteria. Thus, BLAST searching was carried out with the P450 monooxygenase as a probe to identify the candidate BGC for GOMs (gom BGC) from the genome of S. rubellomurinus. Through deletion of the polyketide synthase (PKS) gene in gom BGC and high performance liquid chromatography-mass spectrometry (HPLC-MS) identification of the fermentation products of the mutant strain, the gom BGC can be confirmed to direct the production of GOMs. Furthermore, bioinformatics tools were used to deduce the biosynthetic pathway of GOM-B. [Results] GOM-B and GOM-D were extracted from the fermentation broth of S. rubellomurinus and their structures were identified by NMR. The gom BGC identified in the work was submitted to the PubMed, with the GenBank accession number: OQ831859. The deletion of a PKS gene gomB resulted in the disappearance of GOM-B and GOM-D in the fermentation broth. The biosynthetic pathway of GOM-B was deduced as a type I PKS based on bioinformatics analysis. [Conclusion] This work identified a new BGC which directs the biosynthesis of trialkyl-substituted aromatic polyketides, including GOM-B and GOM-D. The type I PKS involves a P450 monooxygenase GomJ putatively catalyzing the unique polyene chain aromatization. Compared with the gbn BGC recently reported to direct the GOM-G biosynthesis, the PKS assembly line encoded by gom BGC lacks one elongation module, which is consistent with the carbon skeleton of GOM-B. The gom BGC could serve as an example that the gene evolution of bacterial type I PKS leads to the structural diversity of polyketides. Moreover, GomJ shows 78.3% sequence identity to the P450 monooxygenase GbnP functionally identified from gbn BGC, whereas the putative substrate of GomJ has two carbons less than the substrate of GbnP. Thus, the discovery of this work helps to decipher the substrate-specificity mechanism of the unusual P450 monooxygenase which catalyzes polyene chain aromatization in the biosynthetic pathways of trialkyl-substituted aromatic polyketides in bacteria.