PURIFICATION OF CHAPERONINS FROM THERMOPHILIC BACTERIA AND ARCHAEA

Chaperonins are among the most abundant proteins in thermophilic and h yperthermophilic microorganisms. A fast and efficient protocol has bee n designed to purify chaperonins from natural microbial sources on a m illigram scale. The procedure takes advantage of the low isoelectric p oint and high molecular mass of chaperonins. A strong anion-exchange m atrix in combination with gel permeation chromatography and separation on a high-resolution MonoQ column were used to purify four chaperonin s from bacteria and archaea. We also show how the procedure for one of the hyperthermophilic chaperones can be easily scaled up. Pure chaper onins were characterized by two-dimensional polyacrylamide gel electro phoresis and high-performance liquid chromatography to establish heter ogeneity. Pure chaperonins retain the characteristic double ring struc ture in electron microscopy and they bind unfolded proteins.