Search

This paper describes the cloning of a gene (pcp) coding for pyrrolidone carboxylyl peptidase (PYRase), an enzyme which selectively removes N-terminal pyroglutamic acid residues from polypeptides. This gene was isolated from Streptococcus pyogenes by construction of a gene library with a bacteriophage lambda-derived cosmid-Escherichia coli host system. Nucleotide sequence determination of a 1.3 kb restriction fragment revealed a 645 bp open reading frame encoding a 215-amino-acid product of M(r) 23,135 consistent with the 26 kDa polypeptide obtained from in vivo overexpression in E. coli. Southern hybridization confirmed that pcp is a single-copy gene on the S. pyogenes chromosome. 5' and 3' endpoint mapping of the 0.7 kb specific transcript observed by Northern analysis permitted the identification of transcriptional initiation and termination signals. Structural features of the pcp gene product from S. pyogenes are discussed and compared with that from Bacillus subtilis. The lack of sequence identity with any other known protein or nucleotide sequence suggests that this enzyme belongs to a new class of peptidase.

Pyrrolidone carboxyl peptidase (EC 3.4.11.8) (Pcp) is an enzyme that catalyzes the removal of the N-terminal pyroglutamyl group from some peptides or proteins. Its value in protein chemistry and bacterial diagnosis makes this enzyme an interesting subject of study. The present paper reports for the first time the cloning and characterization of a pyrrolidone carboxyl peptidase gene (pcp). This gene is present in a single copy in the genome of Bacillus subtilis as indicated by Southern blot hybridization analysis. The pcp transcripts were analyzed in Escherichia coli by Northern blot hybridization and S1 nuclease mapping. The deduced amino acid sequence predicts a protein of 215 amino acids with a calculated molecular weight of 23,777 Da. The pcp gene has been over-expressed in E. coli, allowing the identification and partial characterization of Pcp protein.