Enhanced Rishirilide Biosynthesis by a Rare In-Cluster Phosphopantetheinyl Transferase in Streptomyces xanthophaeus

Phosphopantetheinyl transferases (PPTases) play important roles in activating apo-acyl carrier proteins (apo-ACPs) and apo-peptidyl carrier proteins (apo-PCPs) in both primary and secondary metabolism. PPTases catalyze the posttranslational modifications of those carrier proteins by covalent attachment of the 49-phosphopantetheine group to a conserved serine residue. The protein-protein interactions between a PPTase and a cognate acyl or peptidyl carrier protein have important regulatory functions in microbial biosynthesis, but the molecular mechanism underlying their specific recognition remains elusive. In this study, we identified a new rishirilide biosynthetic gene cluster with a rare in-cluster PPTase from Streptomyces xanthophaeus no2. The function of this Sfp-type PPTase, SxrX, in rishirilide production was confirmed using genetic mutagenesis and biochemical characterization. We applied molecular modeling and site-directed mutagenesis to identify key residues mediating the protein-protein interaction between SxrX and its cognate ACP. In addition, six natural products were isolated from wild-type S. xanthophaeus no2 and the DsxrX mutant, including rishirilide A and lupinacidin A, that exhibited antimicrobial and anticancer activities, respectively. SxrX is the first Sfptype PPTase identified from an aromatic polyketide biosynthetic gene cluster and shown to be responsible for high-level production of rishirilide derivatives. IMPORTANCE Genome mining has been a vital means for natural product drug discovery in the postgenomic era. The rishirilide-type polyketides have attracted attention due to their potent bioactivity, but the poor robustness of production hosts has limited further research and development. This study not only identifies a hyperproducer of rishirilides but also reveals a rare, in-cluster PPTase SxrX that plays an important role in boosting rishirilide biosynthesis. Experimental and computational investigations revealed new insights on the protein-protein interaction between SxrX and its cognate ACP with wide implications for understanding polyketide biosynthesis.

Automated summary

Phosphopantetheinyl transferases (PPTases) play important roles in activating apo-acyl carrier proteins and apo-peptidyl carrier proteins. This study identified a new rishirilide biosynthetic gene cluster and a rare in-cluster PPTase, SxrX, which plays a crucial role in boosting rishirilide production. Experimental and computational investigations provide insights into the protein-protein interaction between SxrX and its cognate ACP.