The culture temperature affects the properties of the Yersinia pseudotuberculosis porin inclusion bodies and the structure of the recombinant porin

Аннотация

The structure of inclusion bodies (IBs), in addition to the nature of the protein that forms them, depends on a number of parameters of the expression process, including the growth temperature of the cells of the producer strain. In this work we studied the effect of cell cultivation temperature on the properties and structure of Yersinia pseudotuberculosis porin IBs. The inclusion bodies synthesized in E. coli at different temperatures (37, 30 and 18 ° C) were isolated, purified and characterized. Using dynamic light scattering and optical spectroscopy, the solubility of IBs in SDS and urea and the state of the recombinant porin in the denaturants solutions (particle size, degree of polydispersity, secondary structure) were investigated. It was found that lowering the cell growth temperature resulted in a decrease in the stability of IBs in aqueous solutions of urea and SDS and an increase in the proportion of native-like conformations in IBs but also led to an increase in sensitivity of the recombinant porin to chemical denaturation. Mild methods of solubilization of IBs formed at reduced temperature are necessary for the preservation of the native-like porin structure.