Cloning, expression, purification and characterization of a thermo- and surfactant-stable protease from Thermomonospora curvata

Publication date: Available online 26 March 2019Source: Biocatalysis and Agricultural BiotechnologyAuthor(s): Duangjai Sittipol, Pijitra Saelao, Tassanee Lohnoo, Tassanee Lerksuthirat, Yothin Kumsang, Wanta Yingyong, Pongsak Khunrae, Triwit Rattanarojpong, Nujarin JongrujaAbstractProtease is widely used in various industrial applications to hydrolyze the peptide bonds in protein molecules. The appropriate biochemical characteristics of the proteases for the industry, especially thermostability of the enzyme, are the key requirement for the potential applications. Therefore, we identified, expressed, purified and characterized the recombinant protease from thermophilic bacteria, Thermomonospora curvata (Tcsp). We identified the putative gene of serine protease from the genome of T. curvata. Tcsp which without signal peptide was expressed and purified from E. coli as a soluble form. Here, we report that Tcsp worked efficiently at 70 °C and pH 10. The Km for azocasein was 0.94 mg/ml, and Vmax was 605.04 unit/mg. We also show the thermal stability of Tcsp after heat treatment at various temperatures indicating that Tcsp is a thermostable protease. To elucidate whether Tcsp is still active after the pre-incubation with surfactants at high temperature, Tcsp was pre-incubated in the presence of nonionic, cationic, anionic and amphionic surfactants at high temperature to give insight into the applications in the detergent industry. Tcsp shows significant residual activity after ...
Source: Biocatalysis and Agricultural Biotechnology - Category: Biotechnology Source Type: research