The dppA gene of Bacillus subtilis encodes a new D-aminopeptidase

Different strains of Bacillus were screened for their ability to hydrolyse D-alanyl-p-nitroanilide. Activity was detected in Bacillus pumilus, Bacillu s brevis, Bacillus licheniformis 749I and Bacillus subtilis 168. The last s train was the best producer and was selected for the production and purific ation of the enzyme. The determination of the N-terminal sequence identifie d the enzyme as the product of the dppA gene (previously named dciAA) belon ging to the dipeptide ABC transport (dpp) operon expressed early during spo rulation. Open reading frames (ORFs) encoding putative related proteins wer e found in the genomes of a variety of Archaea and both sporulating and non -sporulating bacteria. The enzyme behaves as a D-aminopeptidase and represe nts the prototype of a new peptidase family. Among the tested substrates, t he highest activities were found with D-Ala-d-Ala and D-Ala-Gly-Gly. The ac tive enzyme behaves as an octamer of identical 30 kDa subunits. It exhibits a broad pH optimum, extending between pH 9 and 11. It is reversibly inhibi ted in the presence of Zn2+ chelators, and the sequence comparisons highlig ht the conservation of potential Zn-binding residues. As it has been shown by others that null mutations in the dpp operon do not inhibit spore format ion, the physiological role of DppA is probably an adaptation to nutrient d eficiency.