A Nucleotide-dependent molecular switch controls ATP binding at the C-terminal domain of Hsp90. N-terminal nucleotide binding unmasks a C-terminal binding pocket.
In vivo function of the molecular chaperone Hsp90 is ATP-dependent and requires the full-length protein. Our earlier studies predicted a second C-terminal ATP-binding site in Hsp90. By applying direct biochemical approaches, we mapped two ATP-binding sites and unveiled the C-terminal ATP-binding site as the first example of a cryptic chaperone nucleotide-binding site, which is opened by occupancy of the N-terminal site. We identified an N-terminal gamma-phosphate-binding motif in the middle domain of Hsp90 similar to other GHKL family members. This motif is adjacent to the phosphate-binding region of the C-terminal ATP-binding site. Whereas novobiocin disrupts both C- and N-terminal nucleotide binding, we found a selective C-terminal nucleotide competitor, cisplatin, that strengthens the Hsp90-Hsp70 complex leaving the Hsp90-p23 complex intact. Cisplatin may provide a pharmacological tool to dissect C- and N-terminal nucleotide binding of Hsp90. A model is proposed on the interactions of the two nucleotide-binding domains and the charged region of Hsp90.
Söti
Csaba
C
Department of Medical Chemistry, Semmelweis University, P. O. Box 260, Budapest H-1444, Hungary.
Rácz
Attila
A
Csermely
Péter
P
D000255:Adenosine Triphosphate / Q000378:metabolism* / Q000494:pharmacology; D020816:Amino Acid Motifs; D000595:Amino Acid Sequence; D000818:Animals; D016227:Benzoquinones; D001665:Binding Sites; D002942:Circular Dichroism; D002945:Cisplatin / Q000494:pharmacology*; D003432:Cross-Linking Reagents / Q000494:pharmacology*; D004305:Dose-Response Relationship, Drug; D004591:Electrophoresis, Polyacrylamide Gel; D018840:HSP70 Heat-Shock Proteins / Q000737:chemistry; D018841:HSP90 Heat-Shock Proteins / Q000737:chemistry* / Q000378:metabolism*; D006801:Humans; D019169:Jurkat Cells; D007700:Kinetics; D047029:Lactams, Macrocyclic; D008954:Models, Biological; D008969:Molecular Sequence Data; D009154:Mutation; D011233:Precipitin Tests; D011485:Protein Binding; D017433:Protein Structure, Secondary; D017434:Protein Structure, Tertiary; D011809:Quinones / Q000494:pharmacology; D051381:Rats; D012685:Sepharose / Q000378:metabolism; D017386:Sequence Homology, Amino Acid
D016227:Benzoquinones; D003432:Cross-Linking Reagents; D018840:HSP70 Heat-Shock Proteins; D018841:HSP90 Heat-Shock Proteins; D047029:Lactams, Macrocyclic; D011809:Quinones; D000255:Adenosine Triphosphate; D012685:Sepharose; D002945:Cisplatin; C001277:geldanamycin