Heterologous expression and characterization of a novel subtilisin-like protease from a thermophilic Thermus thermophilus HB8.

Heterologous expression and characterization of a novel subtilisin-like protease from a thermophilic Thermus thermophilus HB8. Int J Biol Macromol. 2019 Jul 16;: Authors: Xie G, Shao Z, Zong L, Li X, Cong D, Huo R Abstract Subtilisins are a family of serine proteases used widely throughout the detergent, leather and food industries, with the identification and development of new enzymes holding much potential value. Thermus thermophilus HB8 was examined for serine proteases and found TTHA0724 gene. Sequence analysis of this putative serine protease placed it within the subtilisin family. To obtain active T. thermophilus HB8 subtilisins, three genes encoding prepro-subtilisin, pro-subtilisin and mature-subtilisin were cloned and expressed in Escherichia coli Transetta (DE3). Although direct expression of the mature-subtilisin gene was found to produce inactive inclusion bodies, expression of the pro-subtilisin gene resulted in active mature-subtilisin, indicating that the pro-sequence of translated pro-subtilisin underwent autoproteolysis. The resulting mature-subtilisin exhibited maximal activity between 65 and 85 °C at pH 7.5. The mature-subtilisin showed good stability, maintaining 50% activity after 48 h at 75 °C and >78% activity across the pH range 5.0-9.5. Furthermore, the mature-subtilisin demonstrated broad substrate specificity, with no requirement for the presence of metal ions which are essential for other sub...
Source: International Journal of Biological Macromolecules - Category: Biochemistry Authors: Tags: Int J Biol Macromol Source Type: research