Discovery of a Phosphonoacetic Acid Derived Natural Product by Pathway Refactoring

Abstract

The activation of silent natural product gene clusters is a synthetic biology problem of great interest. As the rate at which gene clusters are identified outpaces the discovery rate of new molecules, this unknown chemical space is rapidly growing, as too are the rewards for developing technologies to exploit it. One class of natural products that has been underrepresented is phosphonic acids, which have important medical and agricultural uses. Hundreds of phosphonic acid biosynthetic gene clusters have been identified encoding for unknown molecules. Although methods exist to elicit secondary metabolite gene clusters in native hosts, they require the strain to be amenable to genetic manipulation. One method to circumvent this is pathway refactoring, which we implemented in an effort to discover new phosphonic acids from a gene cluster from Streptomyces sp. strain NRRL F-525. By reengineering this cluster for expression in the production host Streptomyces lividans, utility of refactoring is demonstrated with the isolation of a novel phosphonic acid, O-phosphonoacetic acid serine, and the characterization of its biosynthesis. In addition, a new biosynthetic branch point is identified with a phosphonoacetaldehyde dehydrogenase, which was used to identify additional phosphonic acid gene clusters that share phosphonoacetic acid as an intermediate.

Phosphonates: Their Natural Occurrence and Physiological Role

“…An important and instructive example here is an activation of gene cluster from Streptomyces sp. NRRL F-525 and its reengineering in Streptomyces lividans, which resulted in the isolation of O-phosphonoacetic acid serine (compound 42) [91].…”

Refactoring the Cryptic Streptophenazine Biosynthetic Gene Cluster Unites Phenazine, Polyketide, and Nonribosomal Peptide Biochemistry

“…Additional genetic manipulations are often necessary to improve or activate production within heterologous hosts. These may include manipulating the pathway-specific regulatory genes (Olano et al, 2008) or using synthetic biology tools to rewrite native genetic elements with optimized components (e.g., promoters, ribosome binding sites [RBSs]) (Freestone et al, 2017;Montiel et al, 2015;Myronovskyi and Luzhetskyy, 2016).…”

Self-resistance guided genome mining uncovers new topoisomerase inhibitors from myxobacteria

“…Similar pathway-refactoring strategies have been successfully employed for the activation of ‘silent’ gene clusters in Streptomyces and Burkholderia . 24 , 25 Furthermore, promotor insertions have been reported to improve secondary metabolite production by myxobacteria, although the approach did not afford a previously unknown myxobacterial natural product class to date. 2 , 26 …”