Abstract
A phospholipase D (PLD) expression system was constructed from Streptoverticillium cinnamoneum using Escherichia coli as a host. From a sequence alignment analysis of PLD superfamily, a conserved sequence, a GG/S motif, was found, next to the putative active center in the PLD superfamily. Based on the hypothesis that a serine residue in GG/S motif is involved in the catalytic function, a serine residue substituted mutant library (sixteen PLD mutants) was constructed. An analysis of the activities of these mutants showed that two serine residue substituted mutants (G215S and G216S) in the GG motif exhibited high conversion activity. However, the substrate specificity of these mutants differed depending on the donor molecules. When a serine amino acid was used as a donor substrate, G216S showed a high transphosphatidylation activity. For other donors, G215S showed a higher catalytic activity than G216S. These results support the hypothesis that the serine residue in the GG/S motif controls substrate specificity and enhances the turnover of catalytic function.
