Cloning of the Pseudomonas glumae lipase gene and determination of the active site residues.

摘要: The lipA gene encoding the extracellular lipase produced by Pseudomonas glumae PG1 was cloned and characterized. A sequence analysis revealed an open reading frame of 358 codons mature (319 amino acids) preceded a rather long signal 39 acids. As first step in structure-function analysis, we determined Ser-Asp-His triad which makes up catalytic site this lipase. On basis primary homology with other known lipases, number putative active residues located conserved areas were found. To determine actually involved catalysis, constructed substitution mutants for Ser, Asp, His residues. These mutant lipases using P. PG3, from wild-type deleted replacement. By following approach, showed that Ser-87, Asp-241, His-285 make This knowledge, together information on mechanism three-dimensional structure, should facilitate selection specific modifications tailoring industrial applications.