Construction, heterological expression and a simple purification of the BP region of the pneumococcal surface protein A fused in different orientations to the chemotaxis adaptor protein CheW from Thermotoga petrophila

Abstract

Background

The important challenge to the biotechnology is to find new effective fusion partners that enable to improve solubility, expression, and optimize the subsequent fine purification of the target protein.

Results

The most invariant part of the most immunogenic region of the surface virulence factor A of Streptococcus pneumoniae was selected as a model target protein, while the thermostable chemotaxis polypeptide of W-type from Thermotoga petrophila was used as a fusion partner. The genes encoding fusion variants of these proteins were constructed and cloned into a plasmid vector under the control of the strong bacteriophage T7 transcription regulatory system. Effective Escherichia coli producer strains were obtained, and optimal conditions were chosen for the production of resulting constructs. The optimal pH and temperature ranges of recombinant proteins were determined, and three-dimensional shapes of their molecules were also predicted. Methods of low-stage protein purification were improved. Some of the isolated proteins demonstrated a high level of expression, solubility and purity.

Conclusions

Novel chimeric proteins were obtained which had not previously been observed in nature in such domain combinations. It was shown that the biotechnologically valuable characteristics of the hybrid proteins were more marked when the thermal-resistant partner was combined with the N-terminus of pneumococcal protein. The principles of their low-stage purification were performed which does not require any special equipment. That is a basis for significant reduction of prices for diagnostic test-systems components and subunit vaccine production.

How to cite: Grishin DV, Sidorov NG, Parfenova OK, et al. Construction, heterological expression and a simple purification of the BP region of the pneumococcal surface protein a fused in different orientations to the chemotaxis adaptor protein CheW from Thermotoga petrophila. Electron J Biotechnol 2024;71. https://doi.org/10.1016/j.ejbt.2024.05.001.