PURIFICATION AND CHARACTERIZATION OF A DIPEPTIDASE FROM LACTOBACILLUS-HELVETICUS SBT-2171

A metal-dependent dipeptidase was purified to homogeneity from a cell extract of Lactobacillus helveticus SET 2171 by fast protein liquid ch romatography, The enzyme was purified 237-fold from the extract, with a yield of 1.8%, Sodium dodecyl sulfate-polyacrylamide gel electrophor esis of the purified enzyme showed a single protein band with a molecu lar weight of 50,000, The dipeptidase hydrolyzes a range of only dipep tides. Dipeptides containing proline, glutamic acid, and aspartic acid are not hydrolyzed. The enzyme was shown to be a metalloenzyme with a pH optimum of 8.0 and a temperature optimum of 55 degrees C, Dithiol- reducing reagents exert strong inhibition on enzyme activity. Kinetic studies indicated that the enzyme has a relative average affinity for leucyl-leucine (K-m, 0.5 mM), The negative immunoresponse of the purif ied enzyme with monoclonal antibodies raised against a dipeptidase hom Lactococcus lactis subsp, cremoris Wg2 shows that both enzymes can be immunologically distinguished.