摘要：Chaperonin and cochaperonin, represented by E. coli GroEL and GroES, are essential molecular chaperones for protein folding. The double-ring assembly of GroEL is required to function with GroES, and a single-ring GroEL variant GroEL(SR) forms a stable complex with GroES, arresting the chaperoning reaction cycle. GroES I25 interacts with GroEL; however, mutations of I25 abolish GroES-GroEL interaction due to the seven-fold mutational amplification in heptameric GroES. To weaken GroEL(SR)-GroES interaction in a controlled manner, we used groES (7), a gene linking seven copies of groES, to incorporate I25 mutations in selected GroES modules in GroES(7). We generated GroES(7) variants with different numbers of GroESI25A or GroESI25D modules and different arrangements of the mutated modules, and biochemically characterized their interactions with GroEL(SR). GroES(7) variants with two mutated modules participated in GroEL(SR)-mediated protein folding in vitro. GroES(7) variants with two or three mutated modules collaborated with GroEL(SR) to perform chaperone function in vivo: three GroES(7) variants functioned with GroEL(SR) under both normal and heat-shock conditions. Our studies on functional single-ring bacterial chaperonin systems are informative to the single-ring human mitochondrial chaperonin mtHsp60-mtHsp10, and will provide insights into how the double-ring bacterial system has evolved to the single-ring mtHsp60-mtHsp10.
