Although the assembly of multicomponent protein complexes within cells is thought, in many cases, to occur by auto-assembly processes, there is evidence that this is not universally true. Recently, members of a class of proteins known collectively as heat shock proteins have been implicated as playing roles as catalysts mediating a wide variety of protein-protein interactions. The catalytic role these proteins are believed to play is one in which they stabilize incompletely folded proteins thereby allowing them to be further processed or assembled into higher order structures, or they catalyze the unfolding of improperly folded proteins allowing them to regain function. For this reason they have been dubbed 'chaperons.' Recent experimental evidence has been presented showing that certain 'chaperons' directly interact with incompletely folded protein precursors and facilitate their proper folding and higher order assembly. However, the mechanism(s) whereby such processes are mediated is completely unknown. We have recently identified and purified an evolutionarily conserved heat shock protein found in yeast mitochondria and have cloned the gene coding for it. From genetic studies we have found that a mutation in the gene coding for this protein which abolishes the quaternary structure of this protein simultaneously causes the loss of a wide variety of macromolecular assembly processes within the mitochondrial matrix. This evidence is some of the strongest yet which directly implicates a particular heat shock protein as a facilitator of macromolecular assembly. Our goal is to describe the mechanism(s) by which this protein facilitates the stabilization of incompletely folded proteins and catalyzes macromolecular assembly. To achieve this we have chosen to 1) generate a large array of mutants in yeast with altered genes coding for this protein and to characterize their phenotypic effects, and 2) characterize in vitro the properties which this protein possesses when interacting with incompletely or improperly folded polypeptides. Studying the regulation of the mechanism of macromolecular assembly is a relatively new area. The proposed work will provide data on how one particular protein functions in mediating this phenomenon of facilitated assembly.
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