Histidine and serine roles in catalytic activity of choline oxidase from Alcaligenes species studied by chemical modifications

Abstract

This report concentrated on clarifying the histidine and serine roles in choline oxidase (ChOx) biocatalytic activity using chemical modifications. The serine and histidine residues on ChOx were chemically modified by diethylpyrocarbonate (DEPC) and phenylmethanesulfonyl fluoride (PMSF) at 100 mM Tris–HCl buffer, pH 8. The time course, inhibition kinetics and conformational changes of ChOx were investigated using spectrophotometric, fluorescence and Far-UV CD techniques. Enzyme activity decreased of 90% and 50% in the presence of DEPC and PMSF, respectively. The double reciprocal lineweaver-burk plots indicated a competitive and an uncompetitive inhibition pattern for DEPC and PMSF, respectively suggesting the location of histidine in the enzyme active site and that of serine in the outside of active site, but somewhere close to it.