PURIFICATION AND PARTIAL CHARACTERIZATION OF A TRIPEPTIDASE FROM PEDIOCOCCUS-PENTOSACEUS K9.2

A tripeptidase was purified from the cytoplasm of Pediococcus pentosac eus K9.2 by anion-exchange chromatography, gel filtration chromatograp hy, and high-performance liquid chromatography. The molecular mass of the enzyme was estimated by gel filtration at 100,000 Da. Sodium dodec yl sulfate-polyacrylamide gel electrophoresis of the purified peptidas e showed one protein band of 45,000 Da. Optimal enzyme activity was ob tained at pH 7.0 and at 50 degrees C. The peptidase hydrolyzed all tri peptides tested. Cleavage was not observed with dipeptides, oligopepti des, or amino acid-p-nitroanilide derivatives. Strong inhibition of ac tivity was caused by EDTA, 1,10-phenanthroline, dithiothreitol, and be ta-mercaptoethanol, whereas phenylmethylsulfonyl fluoride and sulfur-r eactive reagents had no effect on peptidase activity. Mg2+, Mn2+, and Ca2+ stimulated the hydrolyzing activity of the enzyme. The 20 N-termi nal amino acids of the tripeptidase from P. pentosaceus had 84% identi ty with those from the corresponding N-terminal region of the tripepti dase from Lactococcus lactis subsp. cremoris Wg2.