Structure-Function Studies on Small Heat Shock Protein Oligomeric Assembly and Interaction with Unfolded Polypeptides

This is the authors' abstract. We don't add key points for this paper.

The small heat shock protein (smHSP) and α-crystallin genes encode a family of 12–43-kDa proteins which assemble into large multimeric structures, function as chaperones by preventing protein aggregation, and contain a conserved region termed the α-crystallin domain. Here we report on the structural and functional characterization of Caenorhabditis elegansHSP16-2, a 16-kDa smHSP produced only under stress conditions. A combination of sedimentation velocity, size exclusion chromatography, and cross-linking analyses on wild-type HSP16-2 and five derivatives demonstrate that the N-terminal domain but not most of the the C-terminal extension which follows the α-crystallin domain is essential for the oligomerization of the smHSP into high molecular weight complexes. The N terminus of HSP16-2 is found to be buried within complexes which can accommodate at least an additional 4-kDa of heterologous sequence per subunit. Studies on the interaction of HSP16-2 with fluorescently-labeled and radiolabeled actin and tubulin reveal that this smHSP possesses a high affinity for unfolded intermediates which form early on the aggregation pathway, but has no apparent substrate specificity. Furthermore, both wild-type and C-terminally-truncated HSP16-2 can function as molecular chaperones by suppressing the thermally-induced aggregation of citrate synthase. Taken together, our data on HSP16-2 and a unique 12.6-kDa smHSP we have recently characterized demonstrate that multimerization is a prerequisite for the interaction of smHSPs with unfolded protein as well as for chaperone activity.