Ma. Sentandreu et F. Toldra, BIOCHEMICAL-PROPERTIES OF DIPEPTIDYL PEPTIDASE-III PURIFIED FROM PORCINE SKELETAL-MUSCLE, Journal of agricultural and food chemistry, 46(10), 1998, pp. 3977-3984
Dipeptidyl peptidase III (EC 3.4.14.4) was purified from porcine skele
tal muscle by ammonium sulfate fractionation and by chromatography and
rechromatography on a Biosept-DEAE HPLC column. The enzyme was purifi
ed 2329-fold with a 19% recovery. The relative molecular mass was esti
mated to be 82 000 Da by SDS-PAGE and the maximum activity was reached
at pH 8.0 and 40 degrees C. The enzyme had a wider range of hydrolysi
s on peptide substrates than over dipeptidyl-AMC derivatives, but with
out any endopeptidase activity. On the basis of the kinetic parameters
and the substrates studied, the tetrapeptide Gly-Gly-Phe-Leu (des-Tyr
1 Leu-enkephalin) showed the best affinity for DPP III. The serine pep
tidase inhibitor 3,4-DCI, along with the cysteine protease inhibitors
p-CMB and DTNB were potent inhibitors of DPP III. Chelating and reduci
ng agents also affected considerably the enzyme activity. Co2+ was pro
ved to markedly increase the enzyme activity. Zn2+, Fe2+, Cu2+, Cd2+,
and Hg2+ inhibited the activity, while Ca2+ and Mg2+ had no effect. Th
e dipeptide Tyr-Tyr exerted an important inhibitory effect on DPP III
activity.