Finding key residues for site-directed mutagenesis of haemolytic phospholipase C

FORRELLAD, MA; BEASSONI, PR; DOMENECH, CE; LISA, AT

Pinamar, Buenos Aires

Congreso; XLI Reunión Anual, Sociedad Argentina de Investigación en Bioquímica y Biología Molecular (SAIB); 2005

Sociedad Argentina de Investigación en Bioquímica y Biología Molecular (SAIB)

P. aeruginosa hemolytic phospholipase C (PlcH) is a pathogenic factor that together with PChP supplies Pi and choline. To determine the residues involved in catalysis, bioinfomatic analyses were performed. Sequence profile using BLASTp and CDD revealed that PlcH architecture consists in a highly conserved phosphoesterase domain at N-terminus between residues 51 and 454. This domain is shared with eukariotic acid phosphatases and Gram-negative bacterial Plc.  None of these family members have a resolved structure. Neither sequence nor structure homology with bacterial Gram-positive Plc was observed. LOOPP fold recognition method allowed to identify structural homologues into alkaline phosphatase-like superfamily. The best ranked proteins were E. coli and human alkaline phosphatases. Considering that these enzymes share ligands like Pi and Zn2+, two models of phosphoesterase domain were built to compare the key residues described in these phosphatases. These bioinformatic studies will be useful to perform site-directed mutagenesis to know the role of the conserved aminoacyl residues in the phosphoesterase domain of PlcH