Identification of active sites in amidase: evolutionary relationship between amide bond- and peptide bond-cleaving enzymes.

ABSTRACT: Mainly based on various inhibitor studies previously performed, amidases came to be regarded as sulfhydryl enzymes. Not completely satisfied with this generally accepted interpretation, we performed a series of site-directed mutagenesis studies on one particular amidase of Rhodococcus rhodochrous J1 that was involved in its nitrile metabolism. For these experiments, the recombinant amidase was produced as the inclusion body in Escherichia coli to greatly facilitate its recovery and subsequent purification. With regard to the presumptive active site residue Cys203, a Cys203 --> Ala mutant enzyme still retained 11.5% of the original specific activity. In sharp contrast, substitutions in certain other positions in the neighborhood of Cys203 had a far more dramatic effect on the amidase. Gluta

SUBMITTER: Kobayashi M 

PROVIDER: S-EPMC23678 | biostudies-literature | 1997 Oct

REPOSITORIES: biostudies-literature