Inventaire des différentes activités peptidasiques intracellulaires de Streptococcus thermophilus: Purification et propriétés d'une dipeptide-hydrolase et d'une aminopeptidase

Abstract

Summary Besides a proteolytic activity, four peptidases were isolated from S. thermophilus. The purified dipeptidase has a molecular weight of about 50 000 daltons. It was maximally active around pH 7.50 and 50°C, its apparent energy of activation was 4 500 cal./mole. The dipeptidase was most stable between pH 7.0 and 9.0, and for temperatures less than or equal to 50°C. E.D.T.A. or O-phenanthroline induced a very significant reduction in enzyme activity, but metal ions such as Co++, Zn++, Mn++, Mg++ or Ca++ can reactivate it partially after the inactivation by a chelating agent such as E.D.T.A. Iodoacetate, p-C.M.B. or D.F.P. induced a slight reduction in enzyme activity. Unsubstituted dipeptides only were hydrolysed by the enzyme. The dipeptidase exhibited a specificity toward dipeptides with a large and hydrophobic amino acid residue at the NH2-terminal end; the second amino acid residue (at the COOH-terminal end) had also an effect on enzymic activity. A homogeneous aminopeptidase — with regard to disc electrophoresis — was obtained. The molecular weight of this enzyme was found to be 62 000 daltons. It was maximally active around pH 6.40 and 35°C; its apparent energy of activation was 7 300 cal./mole. Aminopeptidase was most stable between pH 5.80 and 9.80 and at a temperature less than 40°C. E.D.T.A. or O-phenanthroline induced a complete inactivation of the enzyme, but metal ions such as Co++ and Mn++ can reactivate it completely; metal ions such as Ca++ and Zn++ or Mg++ partially after the inactivation by a chelating agent such as E.D.T.A. Zn++ 5.10−3 M and alcohols induced a significant reduction in enzyme activity. This enzyme can be precisely defined as an α-amino-acyl-peptide hydrolase EC.3.4.1. since only NH2-terminal end amino-acid residues were released from oligopeptides (comprising less than eleven amino-acid residues). Leucine-p-nitroanilide and amino-acid amides (except phenylalanine amide) were hydrolysed. Lys-Tyr and Leu-Leu were the best dipeptide substrates. Dipeptides with a glycyl or a histidyl residue at the NH2-terminal end were not hydrolysed as also dipeptides with a prolyl or phenylalanyl residue.