Identification of the peptide binding domain of hsc70. 18-Kilodalton fragment located immediately after ATPase domain is sufficient for high affinity binding.

Abstract

Recombinant hsc70, a purified glutathione S-transferase (GST) fusion protein containing the C-terminal domain of hsc70 (GST-Ct), and an internal 18-kDa polypeptide located immediately after the 44-kDa ATPase domain of hsc70 were investigated for their peptide binding properties. The dissociation constants of the S-peptide for native hsc70 (Kd = 5-8 microM), GST-Ct (Kd = 6.5 microM), and the 18-kDa fragment (Kd = 8.1 microM) are virtually identical. In addition, polylysine and (Pro-Pro-Gly)5 do not show high affinity toward hsc70, GST-Ct, and the 18-kDa fragment, whereas peptide GT4 and P3a show comparably high affinity toward these polypeptides. These observations indicate that the peptide binding domain of hsc70 is confined in the internal 18-kDa fragment.