期刊
ACS CHEMICAL BIOLOGY
卷 15, 期 7, 页码 1813-1819出版社
AMER CHEMICAL SOC
DOI: 10.1021/acschembio.0c00411
关键词
-
资金
- National Institutes of Health [GM116957, GM136235]
Nonribosomal peptide synthetases (NRPSs) are remarkable modular enzymes that synthesize peptide natural products. The condensation (C) domain catalyzes the key amide bond-forming reaction, but structural characterization with bound donor and acceptor substrates has proven elusive. We describe the chemoenzymatic synthesis of condensation domain probes C1 and C2 designed to cross-link the donor and acceptor substrates within the condensation domain active site. These pantetheine probes contain non-hydrolyzable ketone and alpha,alpha-difluoroketone isosteres of the native thioester linkage. Using the bimodular NRPS responsible for synthesis of the siderophore enterobactin as a model system, probe C2 was shown by surface plasmon resonance (SPR) to stabilize an intermolecular interaction between the peptidyl carrier protein (PCP) and C domains in EntB and EntF, respectively, with a dissociation constant of 1-2 nM, whereas the unmodified holo-EntB showed no interaction with EntF. The described condensation domain chemical probes provide powerful tools to study dynamic multifunctional NRPS systems.
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