Identification of the histidine and aspartic acid residues essential for enzymatic activity of a family I.3 lipase by site‐directed mutagenesis

Publisher Website
Google Scholar
Abstract
A lipase from Pseudomonas sp. MIS38 (PML) is a member of the lipase family I.3. We analyzed the roles of the five histidine residues (His30, His274, His291, His313, and His365) and five acidic amino acid residues (Glu253, Asp255, Asp262, Asp275, and Asp290), which are fully conserved in the amino acid sequences of family I.3 lipases, by site-directed mutagenesis. We showed that the mutation of His313 or Asp255 to Ala almost fully inactivated the enzyme, whereas the mutations of other residues to Ala did not seriously affect the enzymatic activity. Measurement of the far- and near-UV circular dichroism spectra suggests that inactivation by the mutation of His313 or Asp255 is not due to marked changes in the tertiary structure. We propose that His313 and Asp255, together with Ser207, form a catalytic triad in PML.