J Biol Chem, Vol. 273, Issue 13, 7351-7357, March 27, 1998

Role of the GroEL Chaperonin Intermediate Domain in Coupling ATP Hydrolysis to Polypeptide Release

From the Department of Molecular Biology, Cell Biology and Biochemistry, Brown University, Box G-J2, Providence, Rhode Island 02912

Modification of the Escherichia coli chaperonin GroEL with N-ethylmaleimide at residue Cys¹³⁸ affects the structural and functional integrity of the complex. Nucleotide affinity and ATPase activity of the modified chaperonin are increased, whereas cooperativity of ATP hydrolysis and affinity for GroES are reduced. As a consequence, release and folding of substrate proteins are strongly impaired and uncoupled from ATP hydrolysis in a temperature-dependent manner. Folding of dihydrofolate reductase at 25 °C becomes dependent on GroES, whereas folding of typically GroES-dependent proteins is blocked completely. At 37 °C, GroES binding is restored to normal levels, and the modified GroEL regains its chaperone activity to some extent. These results assign a central role to the intermediate GroEL domain for transmitting conformational changes between apical and central domains, and for coupling ATP hydrolysis to productive protein release.