Expanding Substrate Promiscuity by Engineering a Novel Adenylating‐Methylating NRPS Bifunctional Enzyme |
| |
Authors: | Dr Sanjib K Shrestha Dr Sylvie Garneau‐Tsodikova |
| |
Affiliation: | Department of Pharmaceutical Sciences, University of Kentucky, BioPharm Complex (Room 423), Lexington, KY, USA |
| |
Abstract: | Nonribosomal peptides synthetases (NRPSs), which are multifunctional mega‐enzymes producing many biologically active metabolites, are ideal targets for enzyme engineering. NRPS adenylation domains play a critical role in selecting/activating the amino acids to be transferred to downstream NRPS domains in the biosynthesis of natural products. Both monofunctional and bifunctional A domains interrupted with an auxiliary domain are found in nature. Here, we show that a bifunctional interrupted A domain can be uninterrupted by deleting its methyltransferase auxiliary domain portion to make an active monofunctional enzyme. We also demonstrate that a portion of an auxiliary domain with almost no sequence identity to the original auxiliary domain can be insert into naturally interrupted A domain to develop a new active bifunctional A domain with increased substrate profile. This work shows promise for the creation of new interrupted A domains in engineered NRPS enzymes. |
| |
Keywords: | domain swapping enzyme catalysis kutzneride natural products nonribosomal peptides thiocoraline |
|
|