Transpeptidase Activity of Streptomyces D-Alanyl-D Carboxypeptidases

Abstract

In the presence of N^α,N^ε-diacetyl-L-Lys-D-Ala-D-Ala as donor, and either D-[¹⁴C]alanine, [¹⁴C]-glycine, or meso-[³H]diaminopimelic acid as acceptor, the DD carboxypeptidases from Streptomyces R61 and R39 catalyze a transpeptidation reaction with the release of terminal D-alanine from the donor and the formation of either N^α,N^ε-diacetyl-L-Lys-D-Ala-D-[¹⁴C]Ala, N^α,N^ε-diacetyl-L-Lys-D-Ala-[¹⁴C] Gly, or N^α,N^ε-diacetyl-L-Lys-D-Ala-D-meso- [³H]diaminopimelic acid. The reaction appears to be a true transpeptidation, and is not simply a “reversal of hydrolysis”. Transpeptidation is inhibited by pencillin at concentrations that inhibit hydrolysis (carboxypeptidase action) of the donor peptide. There are differences in the specificity profiles of the Streptomyces enzymes for acceptor molecules:only the R61 enzyme used [¹⁴C]Gly-Gly as acceptor; transfer of N^α,N^ε-diacetyl-L-Lys-D-Ala to this acceptor resulted in the formation of N^α,N^ε-diacetyl-Lys-D-Ala-[¹⁴C] Gly-Gly, with the synthesis of a (D-Ala-Gly) peptide bond in an endoposition.