1991 Volume 110 Issue 6 Pages 873-878
The prolyl endopeptidase [EC 3. 4. 21. 26] gene of Flavobacterium meningosepticum was cloned in Escherichia coli with the aid of an oligonucleotide probe which was prepared based on the amino acid sequence. The hybrid plasmid, pFPEP1, with a 3. 5 kbp insert at the HincII site of pUC19 containing the enzyme gene, was subcloned into pUC19 to construct plasmid pFPEP3. The whole nucleotide sequence of an inserted HincII-BamHI fragment of plasmid pFPEP3 was determined by the dideoxy chain-terminating method. The purified proly1 endopeptidase was labeled with tritium DFP, and the sequence surrounding the reactive serine residue was found to be Ala (551)-Leu-Ser-Gly-Arg-*Ser-Asn(557). Ser-556 was identified as a reactive serine residue. The enzyme consists of 705 amino acid residues as deduced from the nucleotide sequence and has a molecular weight of 78, 705, which coincides well with the value estimated by ultra centrifugal analysis. The amino acid sequence was 38. 2% homologous to that of the porcine brain prolyl endopeptidase [Rennex et al. (1991) Biochemistry 30, 2195-2203] and 24. 5% homologous to E. coli protease II, which has substrate specificity for basic amino acids [Kanatani et al. (1991) J. Biochem. 110, 315-320].
