Protein folding in the cell - the function of molecular chaperones in facilitating protein folding and protein translocation across membrane
Proteins must fold into their native active conformations to function properly within the cell. A number of genetic diseases, including Huntington's, Alzheimer's, and Creutzfeld Jacob's diseases are caused by defects in protein folding. The work in the Craig lab emphasizes the cellular function of molecular chaperones, a highly conserved group of proteins that facilitates the folding of other proteins by transiently binding to their unfolded forms. Using a combination of experimental tools from biochemistry, genetics, and cell biology, we study cellular processes, including folding of newly synthesized proteins, maintenance of the "prion" form of proteins in cell populations, protein translocation across membranes, and the assembly of Fe/S centers in proteins, in which molecular chaperones play crucial roles. Through analysis of these physiological processes we also aim to understand th ebasis of the functional diversity and specificity of the J-protein/Hsp70 class of proteins that are encoded by large multigene families.
Prion propagation and aggregation of disease-causing proteins
Proteins can exist in more than one conformation. Occasionally protein conformations can be self-propagating and therefore are referred to as prions. Yeast contains several prions whose conformation is profoundly affected by molecular chaperones. In one case, a particular J-protein/Hsp70 chaperone pair is absolutely required for propagation of the prion form with a population of cells. Our goal is to understand the mechanism by which these chaperones affect these self-replicating forms of proteins.
Folding of newly synthesized proteins facilitated by ribosome-associated molecular chaperones
During their synthesis on ribosomes, proteins are particularly susceptible to aggregation, which prohibits their proper folding. We are studying ribosome-associated molecular chaperones that are tethered near the site from which the nascent chain exits the ribosome tunnel. Our goal is to determine how this chaperone and associated factors aid in folding of newly synthesized proteins.
Protein translocation into mitochondria
Hundreds of proteins of the mitochondrial matrix are synthesized on cytosolic ribosomes, then translocated across the mitochondrial inner membrane. The driving force for translocation is a mitochondrial Hsp70. We want to understand the mechanism by which the binding of incoming polypeptide chains by Hsp70 drives import, including the importance of the interaction with Tim44, the membrane associated protein that tethers Ssc1 to the import channel.
Conserved system for assembly of Fe/S clusters and their insertion into proteins
Mitochondria contain a complex system for assembly of Fe-S metal centers and their insertion into proteins. A specialized J-protein/Hsp70 pair is a critical part of this system, interacting specifically with the scafoold protein on which clusters are first built. We aim to unravel the mechanism of action of dedicated chaperone system and the regulation of expression of its components.
For more information see the Craig Lab Website.
Molecular chaperones of the Hsp70 (70) class, working with their co-chaperones Hsp40, facilitate the central cellular processes of protein synthesis, folding degradation, and translocation across membranes. Related, but functionally distinct, Hsp70s function in these different processes (i.e. 70, ribosome associated Hsp70 in protein synthesis and folding of newly synthesized proteins; 70, soluble cytosolic involved in protein folding, degradation, and translocation across membranes; 70, mitochondrial Hsp70 involved in protein translocation and folding).
A prion (an infectious protein) is an altered form of a protein, which can convert the normal (native) form of the protein into the "abnormal" prion form. The prion forms large, ordered aggregates. Fragmentation of these large aggregates increase the number of prion nuclei competent to convert the native form to the prion form, thus increasing the likelihood of transmission of the prion during cell division. Chaperones, including the Hsp70 Ssa and the Hsp40 Sis1, are required for maintenance of certain yeast prions. They may act either by promoting conversion to the prion form or fragmentation of large aggregates.