STRUCTURAL AND FUNCTIONAL-ASPECTS OF BASIC HELIX-LOOP-HELIX PROTEIN-FOLDING BY HEAT-SHOCK PROTEIN-90

The assembly and folding of nascent Polypeptides are mediated in vivo by molecular chaperones, many of which are members of the heat-shock f amily of proteins. We have shown previously that heat-shock protein 90 (HSP90) folds an inactive fraction of a recombinant basic helix-loop- helix (bHLH) protein generated in Escherichia coli (MyoD) into its act ive conformation. We show here that HSP90 also folds another bHLH prot ein (E12) and heterodimers of E12/MyoD into their active conformations . By purifying inactive heterodimers of E12/MyoD and subsequently rend ering them active in binding DNA by treatment with HSP90, we show that one folding step mediated by HSP90 occurs after oligomerization of th e bHLH protein monomers. A series of deletion mutants is used to ident ify the 48-amino acid region of HSP90 that confers bHLH folding activi ty, which lies near the COOH terminus. This region is required for act ivation of DNA binding of MyoD and E12 homodimers and E12/MyoD heterod imers.