Pyrrolidone carboxyl peptidase (EC 3.4.11.8) is an exopeptidase commonly called PYRase, which hydrolytically removes the pGlu-proteins. pGlu also known as pyrrolidone carboxylic acid may occur naturally by an enzymatic procedure or may occur as an artifact in proteins or peptides. The enzymatic synthesis of pGlu suggests that this residue may have important biological and physiological functions. Several studies are consistent with this supposition. PYRase has been found in a variety of bacteria, and in plant, animal, and human tissues. For over two decades, biochemical and enzymatic properties of PYRase have been investigated. At least two classes of PYRase have been characterized. The first one includes the bacterial and animal type I PYRases and the second one the animal type II and serum PYRases. Enzymes from these two classes present differences in their molecular weight and in their enzymatic properties. Recently, the genes of PYRases from four bacteria have been cloned and characterized, allowing the study of the primary structure of these enzymes, and their over-expression in heterelogous organisms. Comparison of the primary structure of these enzymes revealed striking homologies. Type I PYRases and bacterial PYRases are generally soluble enzymes, whereas type II PYRases are membrane-bound enzymes. PYRase II appears to play as important a physiological role as other neuropeptide degrading enzymes. However, the role of type I and bacterial PYRases remains unclear. The primary application of PYRase has been its utilization for some protein or peptide sequencing. Development of chromogenic substrates for this enzyme has allowed its use in bacterial diagnosis.

Pyrrolidone carboxyl peptidasc (EC 3.4.11.8) (Pep) is an enzyme that catalyzes the removal of the N-terminal pyroglutamyl group from some peptidcs 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 gcnc (pep). This gcnc is present in a single copy in the gcnome of Bucilluv sul~rifir as indicated by Soulhcrn blot hybridization analysis, The pep transcripts were analyzed in Escherfchia coti by Northern blot hybridization and S1 nuclcass mapping. The deduced amino acid sequence predicts a protein of 215 amino acids with a calculated molecular weight of 23,777 Da. The pep gene has been over-expressccl in B coli, allowing the identification and parti41 charactcrieation of Pep prodn.

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.