Characterization of functional domains of the eukaryotic co-chaperone Hip.
The homo-oligomeric Hip protein cooperates with the 70-kDa heat shock cognate Hsc70 in the folding of newly synthesized polypeptide chains and in the conformational regulation of signaling molecules known to interact with Hsc70 and Hsp90. In order to further assess the role of Hip during protein biogenesis, a structure-function analysis of the Hip protein was initiated. By employing the yeast two-hybrid system, the Hsc70-binding site of Hip was mapped to a domain comprising multiple tetratricopeptide repeats and flanking charged alpha-helices. Affinity chromatography confirmed direct interaction of isolated Hip fragments and protein fusions bearing this region with the ATPase domain of Hsc70 in an ATP- and salt-dependent manner. Contact of Hip with the ATPase domain appears to be mediated primarily by the positively charged alpha-helix following the tetratricopeptide repeats. Furthermore, a domain required for homo-oligomerization was identified at the extreme amino terminus of Hip.
Irmer
H
H
Zentrum für Molekulare Biologie, Universität Heidelberg, Im Neuenheimer Feld 282, D-69120 Heidelberg, Germany.
Höhfeld
J
J
D000209:Acute-Phase Proteins / Q000737:chemistry*; D000251:Adenosine Triphosphatases / Q000378:metabolism; D000595:Amino Acid Sequence; D000818:Animals; D000951:Antigens, Neoplasm; D001665:Binding Sites; D014408:Biomarkers, Tumor; D002352:Carrier Proteins / Q000737:chemistry / Q000378:metabolism; D002850:Chromatography, Gel; D016384:Consensus Sequence; D050883:HSC70 Heat-Shock Proteins; D018840:HSP70 Heat-Shock Proteins / Q000378:metabolism; D018841:HSP90 Heat-Shock Proteins / Q000737:chemistry / Q000378:metabolism; D037102:Lectins / Q000737:chemistry*; D037181:Lectins, C-Type; D008969:Molecular Sequence Data; D000073718:Pancreatitis-Associated Proteins; D011487:Protein Conformation; D017433:Protein Structure, Secondary; D051381:Rats; D013329:Structure-Activity Relationship
D000209:Acute-Phase Proteins; D000951:Antigens, Neoplasm; D014408:Biomarkers, Tumor; D002352:Carrier Proteins; D050883:HSC70 Heat-Shock Proteins; D018840:HSP70 Heat-Shock Proteins; D018841:HSP90 Heat-Shock Proteins; C494373:Hspa8 protein, rat; D037102:Lectins; D037181:Lectins, C-Type; D000073718:Pancreatitis-Associated Proteins; D000251:Adenosine Triphosphatases