Steps towards the synthetic biology of polyketide biosynthesis

Abstract

Nature is providing a bountiful pool of valuable secondary metabolites, many of which possess therapeutic properties. However, the discovery of new bioactive secondary metabolites is slowing down, at a time when the rise of multidrug-resistant pathogens and the realization of acute and long-term side effects of widely used drugs lead to an urgent need for new therapeutic agents. Approaches such as synthetic biology are promising to deliver a much-needed boost to secondary metabolite drug development through plug-and-play optimized hosts and refactoring novel or cryptic bacterial gene clusters. Here, we discuss this prospect focusing on one comprehensively studied class of clinically relevant bioactive molecules, the polyketides. Extensive efforts towards optimization and derivatization of compounds via combinatorial biosynthesis and classical engineering have elucidated the modularity, flexibility and promiscuity of polyketide biosynthetic enzymes. Hence, a synthetic biology approach can build upon a solid basis of guidelines and principles, while providing a new perspective towards the discovery and generation of novel and new-to-nature compounds. We discuss the lessons learned from the classical engineering of polyketide synthases and indicate their importance when attempting to engineer biosynthetic pathways using synthetic biology approaches for the introduction of novelty and overexpression of products in a controllable manner.

Keywords: combinatorial biosynthesis; drug discovery; plug-and-play biology; refactoring; secondary metabolites.

Figure 1

Pictorial illustration of 6-DEBS synthase, a modular type I PKS and successful attempts at engineering this megasynthase. (a) The native biosynthetic gene cluster and end product. (b) Summary of engineered cluster variants and their products; alterations are indicated in red. Manipulation of the polyketide scaffold includes: (1) substitution of domains (Oliynyk et al., 1996); (2) feeding with noncanonical substrates (Jacobsen et al., 1997); (3) domain insertion (McDaniel et al., 1997); (4) inactivation of domains (Donadio et al., 1993); and (5) domain deletions (Donadio et al., 1991). The effects of modifications 1–5 to the 6-dEB scaffold are also indicated in red, as are the positions at which engineered post-PKS tailoring modifications can occur. AT, acetyltransferase domain; ACP, acyl carrier protein domain; KS, ketosynthase domain; ER, enoyl reductase domain; DH, dehydratase domain.

Figure 2

Schematic illustration of the four steps of polyketide biosynthesis encoded by a prototypical polyketide biosynthetic gene cluster. Each of these steps offers the potential for end product diversification by evolution or engineering as described in the text.