Studies on the primary structures of the exocellular d-alanyl-d-alanine peptidases of Streptomyces strain R61 and Actinomadura strain R39

Abstract

The $\text{M}{}_{\mathrm{<mtext>r</mtext>}}$ 37 000 d-alanyl-d-alanine peptidase excreted by Streptomyces R61 and the $\text{M}{}_{\mathrm{<mtext>r</mtext>}}$ 530 00 d-alanyl-d-alanine peptidase excreted by Actinomadura R39 are both characterized by a very uneven distribution of the basic ($\text{Arg + Lys}$) amino acid residues. Trypsin degradation of the heat-denatured enzymes generates (1) thirteen soluble peptides which contain from 2 to 28 residues in the case of the R61 enzyme and nineteen soluble peptides which contain 2 to 39 residues in the case of the R39 enzyme; and (2) three large segments or core peptides which, irrespective of the enzymes from which they originate, consist of 50–60, 70–80 and 110–120 residues. About 90% of the basic ($\text{Arg + Lys}$) amino acid residues are recovered in the soluble tryptic peptides. The core peptides represent 62% ($\text{M}{}_{\mathrm{<mtext>r</mtext>}}\text{≈ 23 000}$) and 45% ($\text{M}{}_{\mathrm{<mtext>r</mtext>}}\text{≈ 24 000}$) of the untreated R61 and R39 enzymes, respectively. One 28-residue soluble peptide isolated from the R61 enzyme represents the N-terminal portion of the protein whose sequence has been established. The penicillin attachment site of the R61 enzyme has been located in one of the core peptides. For the R39 enzyme, indirect evidence shows that the penicillin binding site is probably within one of the soluble peptides.