期刊
JOURNAL OF THE AMERICAN CHEMICAL SOCIETY
卷 136, 期 4, 页码 1186-1189出版社
AMER CHEMICAL SOC
DOI: 10.1021/ja410462j
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资金
- National Science Foundation, (RCH) [CHE-1058357]
- Federal funds from the National Cancer Institute [Y1-CO-1020]
- National Institute of General Medical Sciences [Y1-GM-1104]
- U.S. Department of Energy, Basic Energy Sciences, Office of Science [DE-AC02-06CH11357]
- Direct For Mathematical & Physical Scien [1058396] Funding Source: National Science Foundation
- Direct For Mathematical & Physical Scien
- Division Of Chemistry [1435915] Funding Source: National Science Foundation
- Division Of Chemistry [1058396] Funding Source: National Science Foundation
Nitrile hydratase (NHase) catalyzes the hydration of nitriles to their corresponding commercially valuable amides at ambient temperatures and physiological pH. Several reaction mechanisms have been proposed for NHase enzymes; however, the source of the nucleophile remains a mystery. Boronic acids have been shown to be potent inhibitors of numerous hydrolytic enzymes due to the open shell of boron, which allows it to expand from a trigonal planar (sp(2)) form to a tetrahedral form (sp(3)). Therefore, we examined the inhibition of the Co-type NHase from Pseudonocardia thermophila JCM 3095 (PtNHase) by boronic acids via kinetics and X-ray crystallography. Both 1-butaneboronic acid (BuBA) and phenylboronic acid (PBA) function as potent competitive inhibitors of PtNHase. X-ray crystal structures for BuBA and PBA complexed to PtNHase were solved and refined at 1.5, 1.6, and 1.2 A resolution. The resulting PtNHase boronic acid complexes represent a snapshot of reaction intermediates and implicate the cysteine-sulfenic acid ligand as the catalytic nucleophile, a heretofore unknown role for the alpha Cys(113)-OH sulfenic acid ligand. Based on these data, a new mechanism of action for the hydration of nitriles by NHase is presented.
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