Nitrile hydratase (nitrile hydratase, NHase, EC 4.2.1.84) is one type of metalloenzyme that catalyzes the formation of amides from nitriles. It is used in the biological production of acrylamide and nicotinamide in industry. Since the hydration of nitriles is an exothermic reaction, the thermal stability of nitrile hydratase attracted increasing attention. Previously, a novel nitrile hydratase derived from Caldalkalibacillus thermarum TA2.A1 was obtained through gene mining. Cal.t NHase possesses excellent thermal stability, while the enzyme activity is far behind the requirements of industrial production. Therefore, improving catalytic efficiency is important for its further applications. This study adopted molecular dynamics simulation methods and found two key residues, N47 and N181, located at the entrance of the substrate channel on the b subunit. Making representative residues mutations at these sites breaks the hydrogen bond, increases the flexibility of the substrate channel entrance, and then improves the enzyme activity. The optimal mutant, βN47F, compared with the wild-type enzyme, shows a 2.57-fold increase in acrylonitrile catalytic activity, and maintain good thermal stability at the same time. This variant enables the industrial application of Cal.t NHase in the future.
ZHANG Weimiao
,
CHENG Zhongyi
,
ZHOU Li
,
ZHOU Zhemin
. Modification of the key amino residues locating the substrate channel entrance of nitrile hydratase that regulate the enzyme activity[J]. Food and Fermentation Industries, 2022
, 48(9)
: 8
-13
.
DOI: 10.13995/j.cnki.11-1802/ts.029373
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