Evolution-guided engineering of <i>trans</i>-acyltransferase polyketide synthases.

Abstract

Bacterial multimodular polyketide synthases (PKSs) are giant enzymes that generate a wide range of therapeutically important but synthetically challenging natural products. Diversification of polyketide structures can be achieved by engineering these enzymes. However, notwithstanding successes made with textbook <i>cis</i>-acyltransferase (<i>cis</i>-AT) PKSs, tailoring such large assembly lines remains challenging. Unlike textbook PKSs, <i>trans</i>-AT PKSs feature an extraordinary diversity of PKS modules and commonly evolve to form hybrid PKSs. In this study, we analyzed amino acid coevolution to identify a common module site that yields functional PKSs. We used this site to insert and delete diverse PKS parts and create 22 engineered <i>trans</i>-AT PKSs from various pathways and in two bacterial producers. The high success rates of our engineering approach highlight the broader applicability to generate complex designer polyketides.