The chaperonin GroEL is able to mediate protein folding in its central cavity. GroEL-bound dihydrofolate reductase assumes its native conformation when the GroES cofactor caps one end of the GroEL cylinder, thereby discharging the unfolded polypeptide into an enclosed cage. Folded dihydrofolate reductase emerges upon ATP-dependent GroES release. Other proteins, such as rhodanese, may leave GroEL after having attained a conformation that is committed to fold. Incompletely folded polypeptide rebinds to GroEL, resulting in structural rearrangement for another folding trial in the chaperonin cavity.

Various diverse extracellular proteins possess Ca(2+)-binding epidermal growth factor (EGF)-like domains, the function of which remains uncertain. We have determined, at high resolution (1.5 A), the crystal structure of such a domain, from human clotting factor IX, as a complex with Ca2+. The Ca2+ ligands form a classic pentagonal bipyramid with six ligands contributed by one polypeptide chain and the seventh supplied by a neighboring EGF-like domain. The crystal structure identifies the role of Ca2+ in maintaining the conformation of the N-terminal region of the domain, but more importantly demonstrates that Ca2+ can directly mediate protein-protein contacts. The observed crystal packing of the domains provides a plausible model for the association of multiple tandemly linked EGF-like domains in proteins such as fibrillin-1, Notch, and protein S. This model is consistent with the known functional data and suggests a general biological role for these domains.

Heat shock proteins (HSPs) derived from tumors or virally infected cells can stimulate antigen-specific CD8+ T cell responses in vitro and in vivo. Although this antigenicity is known to arise from HSP-associated peptides presented to the immune system by major histocompatibility complex (MHC) class I molecules, the cell biology underlying this presentation process remains poorly understood. Here we show that HSP 70 binds to the surface of antigen presenting cells by a mechanism with the characteristics of a saturable receptor system. After this membrane interaction, processing and MHC class I presentation of the HSP-associated antigen can occur via either a cytosolic (transporter associated with antigen processing [TAP] and proteasome–dependent) or an endosomal (TAP and proteasome–independent) route, with the preferred pathway determined by the sequence context of the optimal antigenic peptide within the HSP-associated material. These findings not only characterize two highly efficient, specific pathways leading to the conversion of HSP-associated antigens into ligands for CD8+ T cells, they also imply the existence of a mechanism for receptor-facilitated transmembrane transport of HSP or HSP-associated ligands from the plasma membrane or lumen of endosomes into the cytosol.

The reaction mechanism of protein folding by the chaperonin GroEL and its regulator GroES has been defined. GroES and substrate protein counteract each other's effects on GroEL: whereas GroES stabilizes GroEL in the ADP-bound state, binding of unfolded polypeptide within the cavity of the GroEL cylinder triggers ADP and GroES release. Upon ADP-ATP exchange, GroES reassociates with GroEL and ATP hydrolysis discharges the bound protein for folding. Partially folded protein rebinds to the chaperonin, thus perpetuating the cycle until folding is complete.