SECONDARY STRUCTURE OF THE MAMMALIAN 70-KILODALTON HEAT-SHOCK COGNATE PROTEIN ANALYZED BY CIRCULAR-DICHROISM SPECTROSCOPY AND SECONDARY STRUCTURE PREDICTION

Heat shock proteins are rapidly synthesized when cells are exposed to stressful agents that cause protein damage. The 70-kDa heat shock induced proteins and their closely related constitutively expressed cognate proteins bind to unfolded and aberrant polypeptides and to hydrophilic peptides. The structural features of the 70-kDa heat shock proteins that confer the ability to associate with diverse polypeptides are unknown. In this study, we have used circular dichroism (CD) spectroscopy and secondary structure prediction to analyze the secondary structure of the mammalian 70-kDa heat shock cognate protein (hsc 70). The far-ultraviolet CD spectrum of hsc 70 indicates a large fraction of α-helix in the protein and resembles the spectra one obtains from proteins of the α/β structural class. Analysis of the CD spectra with deconvolution methods yielded estimates of secondary structure content. The results indicate about 40% α-helix and 20% aperiodic structure within hsc 70 and between 16–41% β-sheet and 21–0% β-turn. The Garnier–Osguthorpe–Robson method of secondary structure prediction was applied to the rat hsc 70 amino acid sequence. The predicted estimates of α-helix and aperiodic structure closely matched the values derived from the CD analysis, whereas the predicted estimates of β-sheet and β-turn were midway between the CD-derived values. Present evidence suggests that the polypeptide ligand binding domain of the 70-kDa heat shock protein resides within the C-terminal 160 amino acids [Milarski, K. L., & Morimoto, R. I. (1989) J. Cell Biol. 109, 1947-1962]. Much of this region in hsc 70 is unusually rich in charged and polar amino acids and is predicted to have a high α-helical content. A model for polypeptide binding based on these structural features is proposed. © 1990, American Chemical Society. All rights reserved.