Proline-specific extracellular aminopeptidase purified from Streptomyces lavendulae

Aminopeptidases catalyze the cleavage of specific amino acids from the amino terminus of protein or peptide substrates. A proline-specific aminopeptidase was purified to homogeneity from the culture-free extract of Streptomyces lavendulae ATCC 14162 in sequential steps comprising ammonium sulfate precipitation, ultra-filtration, and column chromatography on Q-sepharose and Sephadex G-100. The purified protein showed approximately 60 kDa in SDS-PAGE and was optimally active at pH 6.5 and 40 °C. Kinetic studies showed a K(m) and V(max) of 0.23 mM and 0.087 μmol/min, respectively, using Pro-p-NA, the substrate with maximum specificity. Enzyme activity was inhibited by PMSF and ions like Zn²+, Co²+, and Ni²+. However, unlike other aminopeptidases, the activity was enhanced in the presence of DTT, 1,10-phenanthroline, EDTA, amastatin, and bestatin. Ions like Ca²+, Mg²+, and Mn²+ also enhanced the activity.