TY - JOUR
T1 - Identification of the N-terminal Peptide Binding Site of Glucose-regulated Protein 94
AU - Gidalevitz, Tali
AU - Biswas, Chhanda
AU - Ding, Hua
AU - Schneidman-Duhovny, Dina
AU - Wolfson, Haim J.
AU - Stevens, Fred
AU - Radford, Sheena
AU - Argon, Yair
PY - 2004/4/16
Y1 - 2004/4/16
N2 - Because the stress protein GRP94 can augment presentation of peptides to T cells, it is important to define how it, as well as all other HSP90 family members, binds peptides. Having previously shown that the N-terminal half of GRP94 can account for the peptide binding activity of the full-length protein, we now locate this binding site by testing predictions of a molecular docking model. The best predicted site was on the opposite face of the β sheet from the pan-HSP90 radicicol-binding pocket, in close proximity to a deep hydrophobic pocket. The peptide and radicicol-binding sites are distinct, as shown by the ability of a radicicol-refractive mutant to bind peptide. When the fluorophore acrylodan is attached to Cys117 within the hydrophobic pocket, its fluorescence is reduced upon peptide binding, consistent with proximity of the two ligands. Substitution of His125, which contacts the bound peptide, compromises peptide-binding activity. We conclude that peptide binds to the concave face of the β sheet of the N-terminal domain, where binding is regulated during the action cycle of the chaperone.
AB - Because the stress protein GRP94 can augment presentation of peptides to T cells, it is important to define how it, as well as all other HSP90 family members, binds peptides. Having previously shown that the N-terminal half of GRP94 can account for the peptide binding activity of the full-length protein, we now locate this binding site by testing predictions of a molecular docking model. The best predicted site was on the opposite face of the β sheet from the pan-HSP90 radicicol-binding pocket, in close proximity to a deep hydrophobic pocket. The peptide and radicicol-binding sites are distinct, as shown by the ability of a radicicol-refractive mutant to bind peptide. When the fluorophore acrylodan is attached to Cys117 within the hydrophobic pocket, its fluorescence is reduced upon peptide binding, consistent with proximity of the two ligands. Substitution of His125, which contacts the bound peptide, compromises peptide-binding activity. We conclude that peptide binds to the concave face of the β sheet of the N-terminal domain, where binding is regulated during the action cycle of the chaperone.
UR - http://www.scopus.com/inward/record.url?scp=1942469336&partnerID=8YFLogxK
U2 - 10.1074/jbc.M313060200
DO - 10.1074/jbc.M313060200
M3 - Article
C2 - 14754890
AN - SCOPUS:1942469336
SN - 0021-9258
VL - 279
SP - 16543
EP - 16552
JO - Journal of Biological Chemistry
JF - Journal of Biological Chemistry
IS - 16
ER -