Purification and Characterization of Alkaline Serine Protease from an Alkalophilic Streptomyces sp.

SAP, an extracellular alkaline serine protease produced by Streptomyces sp. YSA-130, was purified to homogeneity by CM-Sephadex column chromatography and crystallization. The enzyme was a monomeric protein with a molecular weight of 19, 000 as estimated by SDS–PAGE and gel filtration. The amino acid composition and amino-terminal sequence of SAP were similar to those of other bacterial serine proteases, i.e., Streptomyces griseus proteases A and B, Lysobacter enzymogenes a-lytic protease and Nocardiopsis dassonvillei subsp. prasina OPC-210 alkaline serine protease NDP-I. The optimum temperature and pH for the enzyme activity were 60°C and 11.5. The enzyme was stable up 50°C, and between pHs 4 and 12. The activity was inhibited by Ag +, Hg2 +, Co2 +, sodium dodecyl sulfate, N-bromosuccinimide, diisopropyl phosphorofluoridate (DFP), 2, 3-butanedione, 5, 5′-dithiobis-(2-nitrobenzoic acid) (DTNB), iodoacetate,N-ethylmaleimide (NEM), phenylmethanesulfonyl fluoride (PMSF), and phenylglyoxal.