Aminolytic reaction catalyzed by d-stereospecific amidohydrolases from Streptomyces spp

From investigation of 2000 soil isolates, we identified two serine-type amidohydrolases that can hydrolyze d-aminoacyl derivatives from the culture supernatant of Streptomyces species 82F2 and 83D12. The enzymes, redesignated as 82F2-DAP and 83D12-DAP, were purified for homogeneity and characterized. Each enzyme had molecular mass of approximately 40 kDa, and each showed moderate stability with respect to temperature and pH. Among hydrolytic activities toward d-aminoacyl-pNAs, the enzymes showed strict specificity toward d-Phe-pNA, but showed broad specificity toward d-aminoacyl esters. The specific activity for d-Phe-pNA hydrolysis of 82F2-DAP was ten-fold higher than that of 83D12-DAP. As a second function, each enzyme showed peptide bond formation activity by its function of aminolysis reaction. Based on results of d-Phe–d-Phe synthesis under various conditions, we propose a reaction mechanism for d-Phe–d-Phe production. Furthermore, the enzymes exhibited peptide elongation activity, producing oligo homopeptide in a one-pot reaction. We cloned the genes encoding each enzyme, which revealed that the primary structure of each enzyme showed 30–60% identity with those of peptidases belonging to the clan SE, S12 peptidase family categorized as serine peptidase with d-stereospecificity.

► Streptomyces spp. 82F2 and 83D12 secrete serine-type d-stereospecific amidohydrolases.
► The enzymes show peptide bond formation activity by their function of aminolysis reaction.
► The enzymes exhibit peptide elongation activity, producing oligo homopeptide in a one-pot reaction.
► The enzymes belong to the clan SE, S12 peptidase family.