SINGLE-STAGE PURIFICATION OF MOLECULAR CHAPERONE GroEL BY THE AFFINITY CHROMATOGRAPHY ON A BASIS OF DENATURED PEPSIN

Abstract

GroEL (oligomeric heat-shock protein of Escherichia coli cells) is a representative of a group of molecular chaperones that are able to prevent aggregation of nonnative conformational states of proteins by an interaction with them. In the present work we demonstrate a possibility of fast and high-effective purification of GroEL using the chromatography carrier (sepharose) with nonnative pepsin covalently bound to it. The conditions that allow to produce high-purity native GroEL (on the data of SDS-electrophoresis in polyacrylamide gel, fluorescence spectroscopy and ATP‑ase activity) from cell lysate in one step of purification are presented. The method of affinity chromatography can be used for preparative isolation of GroEL and GroEL-like chaperones from different types of cells as well as for express-analysis of a content of chaperones in cells after the action of different stress or pathogenic factors.