LYTIC ENZYMES OF SORANGIUM SP.: ACTION OF THE α- AND β-LYTIC PROTEASES ON TWO BACTERIAL MUCOPEPTIDES

Abstract

The α- and β-lytic proteases hydrolyzed mucopeptides from the walls of Arthrohacter globiformis and Micrococcus lysodeikticus cells. The Arthrobacter mucopeptide was the more readily degraded substrate; the β-enzyme was the more active enzyme. Hydrolysates of Micrococcus mucopeptide showed the simpler electrophoretic pattern of ninhydrin-positive components.Analyses for C-terminal amino acids (after hydrazinolysis) and for dinitrophenyl amino acids (after dinitrophenylation) showed that the hydrolyses of Micrococcus mucopeptide were accompanied by increases in (a) C-terminal glycine and, to a lesser extent, in (b) C-terminal alanine, and by increases in (c) free ε-amino groups of lysine residues and in (d) N-terminal alanine residues. A brief hydrolysis with the β-enzyme gave slightly to moderately greater increases in a, b, and c, and a much greater increase in d, than a prolonged hydrolysis with the α-enzyme at a higher enzyme concentration.The soluble peptides hydrolyzed from Micrococcus mucopeptide by each enzyme were isolated by ion-exchange chromatography. Their composition, and that of the main hydrolysis product of the α-enzyme (an insoluble gel), was compared with that of untreated mucopeptide. The amino acids of the four products remained in approximately the same molar ratio relative to glutamic acid: Ala_2.2, Gly_1.0, Lys_1.0, Glu_1.0. N-Acetyl muramic acid and N-acetyl glucosamine were estimated by gas–liquid chromatography. Relative to glutamic acid, the molar ratio of each amino sugar was about 3.0 in the mucopeptide, 5.0 in the gel, and 0.6 in the soluble peptides. Half the glycyl residues of the mucopeptide and virtually all the glycyl residues of the other three products were analyzed as C-terminal glycine. Subtractive Edman analyses showed little change in the Ala:Glu ratio of the mucopeptide and of the gel but a substantial decrease in the ratios for the soluble peptides. The titratable acidity of the mucopeptide and the gel was consistent with these analyses.It is concluded that cross-linkages from the C-terminus of the peptide chains of the mucopeptide are hydrolyzed by both enzymes and that the linkage between N-acetyl muramic acid and the alanyl residue at the N-terminus of the peptide chains is hydrolyzed rapidly by the β-enzyme and slowly by the α-enzyme.