HspBP1, an Hsp70 cochaperone, has two structural domains and is capable of altering the conformation of the Hsp70 ATPase domain.
We present here the first structural information for HspBP1, an Hsp70 cochaperone. Using circular dichroism, HspBP1 was determined to be 35% helical. Although HspBP1 is encoded by seven exons, limited proteolysis shows that it has only two structural domains. Domain I, amino acids 1-83, is largely unstructured. Domain II, amino acids 84-359, is predicted to be 43% helical using circular dichroism. Using limited proteolysis we have also shown that HspBP1 association changes the conformation of the ATPase domain of Hsp70. Only domain II of HspBP1 is required to bring about this conformational change. Truncation mutants of HspBP1 were tested for their ability to inhibit the renaturation of luciferase and bind to Hsp70 in reticulocyte lysate. A carboxyl terminal truncation mutant that was slightly longer than domain I was inactive in these assays, but domain II was sufficient to perform both functions. Domain II was less active than full-length HspBP1 in these assays, and addition of amino acids from domain I improved both functions. These studies show that HspBP1 domain II can bind Hsp70, change the conformation of the ATPase domain, and inhibit Hsp70-associated protein folding.
McLellan
Catherine A
CA
Department of Molecular and Cellular Biology, University of Arizona, Tucson, Arizona 85721-0038, USA.
Raynes
Deborah A
DA
Guerriero
Vince
V
D048868:Adaptor Proteins, Signal Transducing; D000251:Adenosine Triphosphatases / Q000737:chemistry*; D000255:Adenosine Triphosphate / Q000378:metabolism; D002352:Carrier Proteins / Q000737:chemistry* / Q000235:genetics / Q000494:pharmacology*; D002918:Chymotrypsin / Q000378:metabolism; D002942:Circular Dichroism; D019286:Endopeptidase K / Q000378:metabolism; D005091:Exons; D018840:HSP70 Heat-Shock Proteins / Q000737:chemistry*; D006801:Humans; D008156:Luciferases / Q000037:antagonists & inhibitors / Q000737:chemistry; D013058:Mass Spectrometry; D016254:Mutagenesis, Insertional; D016297:Mutagenesis, Site-Directed; D011487:Protein Conformation / Q000187:drug effects*; D020673:Protein Renaturation / Q000187:drug effects; D017433:Protein Structure, Secondary; D011994:Recombinant Proteins / Q000378:metabolism; D012156:Reticulocytes; D013329:Structure-Activity Relationship; D014357:Trypsin / Q000378:metabolism
D048868:Adaptor Proteins, Signal Transducing; D002352:Carrier Proteins; D018840:HSP70 Heat-Shock Proteins; C116331:HSPBP1 protein, human; D011994:Recombinant Proteins; D000255:Adenosine Triphosphate; D008156:Luciferases; D002918:Chymotrypsin; D014357:Trypsin; D019286:Endopeptidase K; D000251:Adenosine Triphosphatases