HSC70-BINDING PEPTIDES SELECTED FROM A PHAGE DISPLAY PEPTIDE LIBRARY THAT RESEMBLE ORGANELLAR TARGETING SEQUENCES

A 15-mer phage display random peptide library was screened with purifi ed bovine Hsc70, and nucleotide sequence analysis of the selected clon es showed a large enrichment for peptides containing basic sequences w ith at least KR, RR, or RR, Binding affinity for Hsc70 of representati ve peptides increased dramatically for heptamers compared with hexamer s, The peptide RIVRKKK, had the highest affinity for Hsc70, and substi tution analyses showed that hydrophobic residues followed by basic res idues play important roles in maintaining this affinity, In contrast, NIVRKKK was a weaker stimulator of the Hsc70 ATPase activity compared with pigeon cytochrome c peptide and FYQLALT, a peptide optimized for binding to Hsc70, FYQLALT effectively blocked the binding of NIVRKKK t o Hsc70, possibly by causing a conformational change that masked Hsc70 's binding site for the basic peptide, Two hypotheses are offered to e xplain the two different peptide motifs, First, it is proposed that Hs c70 recognizes two different amino acid sequence motifs in its dual ro les of chaperoning proteins to organelles (NIVRKKK-like sequences) and facilitating protein folding (FYQLALT-like sequences), Second, the NI VRKKK motif may be used to bind certain folded proteins with which Hsc 70 interacts, such as itself, p53, and Dnaj2.