title: Nucleotide-dependent interactions within a specialized Hsp70/Hsp40 complex involved in Fe-S cluster biogenesis.
abstract: The structural mechanism by which Hsp70-type chaperones interact with Hsp40-type co-chaperones has been of great interest, yet still remains a matter of debate. Here, we used solution NMR spectroscopy to investigate the ATP-/ADP-dependent interactions between Escherichia coli HscA and HscB, the specialized Hsp70/Hsp40 molecular chaperones that mediate iron-sulfur cluster transfer. We observed that NMR signals assigned to amino acid residues in the J-domain and its "HPD" motif of HscB broadened severely upon the addition of ATP-bound HscA, but these signals were not similarly broadened by ADP-bound HscA or the isolated nucleotide binding domain of HscA complexed with either ATP or ADP. An HscB variant with an altered HPD motif, HscB(H32A,P33A,D34A), failed to manifest WT-like NMR signal perturbations and also abolished WT-like stimulation of ATP hydrolysis by HscA. In addition, residues 153-171 in the C-terminal region of HscB exhibited NMR signal perturbations upon interaction with HscA, alone or complexed with ADP or ATP. These results demonstrate that the HPD motif in the J-domain of HscB directly interacts with ATP-bound HscA and suggest that a second, less nucleotide-dependent binding site for HscA resides in the C-terminal region of HscB.
authors: Mitochondrial Protein Partnership, Center for Eukaryotic Structural Genomics, and ‡Department of Biochemistry, University of Wisconsin , Madison, Wisconsin 53706, United States.
authors: Jin Hae
authors: JH
authors: Kim
authors: T Reid
authors: TR
authors: Alderson
authors: Ronnie O
authors: RO
authors: Frederick
authors: John L
authors: JL
authors: Markley
mesh_terms: D000244:Adenosine Diphosphate / Q000737:chemistry; D000255:Adenosine Triphosphate / Q000737:chemistry; D004926:Escherichia coli / Q000737:chemistry; D050956:HSP40 Heat-Shock Proteins / Q000737:chemistry / Q000378:metabolism*; D018840:HSP70 Heat-Shock Proteins / Q000737:chemistry / Q000378:metabolism*; D007506:Iron-Sulfur Proteins / Q000096:biosynthesis* / Q000737:chemistry*; D019906:Nuclear Magnetic Resonance, Biomolecular; D009711:Nucleotides / Q000737:chemistry* / Q000378:metabolism*
chemical_list: D050956:HSP40 Heat-Shock Proteins; D018840:HSP70 Heat-Shock Proteins; D007506:Iron-Sulfur Proteins; D009711:Nucleotides; D000244:Adenosine Diphosphate; D000255:Adenosine Triphosphate