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 family 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 active conformation. We show here that HSP90 also folds another bHLH protein (E12) and heterodimers of E12/MyoD into their active conformations. By purifying inactive heterodimers of E12/MyoD and subsequently rendering them active in binding DNA by treatment with HSP90, we show that one folding step mediated by HSP90 occurs after oligomerization of the bHLH protein monomers. A series of deletion mutants is used to identify the 48-amino acid region of HSP90 that confers bHLH folding activity, which lies near the COOH terminus. This region is required for activation of DNA binding of MyoD and E12 homodimers and E12/MyoD heterodimers.
Shue
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Department of Pathology, Mount Sinai School of Medicine, New York, New York 10029.
Kohtz
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D000595:Amino Acid Sequence; D000818:Animals; D003001:Cloning, Molecular; D017124:Conserved Sequence; D004330:Drosophila / Q000378:metabolism; D004926:Escherichia coli; D006360:Heat-Shock Proteins / Q000378:metabolism*; D018257:Helix-Loop-Helix Motifs; D046911:Macromolecular Substances; D051379:Mice; D008969:Molecular Sequence Data; D016296:Mutagenesis; D017570:MyoD Protein / Q000096:biosynthesis / Q000737:chemistry* / Q000378:metabolism; D017510:Protein Folding; D051381:Rats; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis / Q000302:isolation & purification; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000737:chemistry / Q000378:metabolism; D017384:Sequence Deletion; D017386:Sequence Homology, Amino Acid; D014345:Trypanosoma / Q000378:metabolism
D006360:Heat-Shock Proteins; D046911:Macromolecular Substances; D017570:MyoD Protein; D011993:Recombinant Fusion Proteins; D011994:Recombinant Proteins