Structures of unliganded and ATP-bound states of the Escherichia coli chaperonin GroEL by cryoelectron microscopy

We have developed an angular refinement procedure incorporating correction for the microscope contrast transfer function, to determine cryoelectron mi croscopy (cryo-EM) structures of the Escherichia coli chaperonin GroEL in i ts apo and ATP-bound forms. This image reconstruction procedure is verified to 13-Angstrom resolution by comparison of the cryo-EM structure of unliga nded GroEL with the crystal structure. Binding, encapsulation, and release of nonnative proteins by GroEL and its cochaperone GroES are controlled by the binding and hydrolysis of ATP. Seven ATP molecules bind cooperatively t o one heptameric ring of GroEL. This binding causes long-range conformation al changes that determine the orientations of remote substrate-binding site s, and it also determines the conformation of subunits in the opposite ring of GroEL, in a negatively cooperative mechanism. The conformation of GroEL -ATP was determined at similar to 15-Angstrom resolution. In one ring of Gr oEL-ATP, the apical (substrate-binding) domains are extremely disordered, c onsistent with the high mobility needed for them to achieve the 60 degrees elevation and 90 degrees twist of the GroES-bound state. Unexpectedly, ATP binding also increases the separation between the two rings, although the i nterring contacts are present in the density map. (C) 2001 Academic Press.