ISOLATION AND CHARACTERIZATION OF PCP, A GENE ENCODING A PYRROLIDONE CARBOXYL PEPTIDASE IN STAPHYLOCOCCUS-AUREUS

The pcp gene, encoding a pyrrolidone carboxyl peptidase (PYRase), was cloned from a lambda GT11 genomic library prepared from Staphylococcus aureus FDA 574 and sequenced. The pcp gene is located 740 bp downstre am from cna, a gene that encodes a collagen-binding adhesin in S. awur eus S. aureus pcp encodes a 212-amino-acid (aa) polypeptide. The pcp g ene was overexpressed in Escherichia coli and the PYRase purified to h omogeneity. The recombinant enzyme exhibited biological activitity, as determined using the chromogenic substrate L-pyroglutamyl-beta-napthy lamide. Biochemical analysis of the PYRase using thiol-blocking chemic als suggested that the enyzme belongs to the cysteine peptidase family . Moreover, multiple sequence alignment revealed a high degree of simi larity to previously described bacterial PYRases. This family of pepti dases has been used to selectively remove the N-terminal pyrrolidone c arboxylic acid residue found on certain blocked proteins and peptides prior to aa sequencing. However, the exact biological role of PYRases has yet to be elucidated.