In:
Proceedings of the National Academy of Sciences, Proceedings of the National Academy of Sciences, Vol. 110, No. 8 ( 2013-02-19), p. 2810-2815
Abstract:
Aldoxime dehydratase (OxdA), which is a unique heme protein, catalyzes the dehydration of an aldoxime to a nitrile even in the presence of water in the reaction mixture. Unlike the utilization of H 2 O 2 or O 2 as a mediator of catalysis by other heme-containing enzymes (e.g., P450), OxdA is notable for the direct binding of a substrate to the heme iron. Here, we determined the crystal structure of OxdA. We then constructed OxdA mutants in which each of the polar amino acids lying within ∼6 Å of the iron atom of the heme was converted to alanine. Among the purified mutant OxdAs, S219A had completely lost and R178A exhibited a reduction in the activity. Together with this finding, the crystal structural analysis of OxdA and spectroscopic and electrostatic potential analyses of the wild-type and mutant OxdAs suggest that S219 plays a key role in the catalysis, forming a hydrogen bond with the substrate. Based on the spatial arrangement of the OxdA active site and the results of a series of mutagenesis experiments, we propose the detailed catalytic mechanism of general aldoxime dehydratases: ( i ) S219 stabilizes the hydroxy group of the substrate to increase its basicity; ( ii ) H320 acts as an acid-base catalyst; and ( iii ) R178 stabilizes the heme, and would donate a proton to and accept one from H320.
Type of Medium:
Online Resource
ISSN:
0027-8424
,
1091-6490
DOI:
10.1073/pnas.1200338110
Language:
English
Publisher:
Proceedings of the National Academy of Sciences
Publication Date:
2013
detail.hit.zdb_id:
209104-5
detail.hit.zdb_id:
1461794-8
SSG:
11
SSG:
12
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