title: Biological heterogeneity of the peptide-binding motif of the 70-kDa heat shock protein by surface plasmon resonance analysis.
abstract: 70-kDa heat shock protein family is a molecular chaperone that binds to a variety of client proteins and peptides in the cytoplasm. Several studies have revealed binding motifs between 70-kDa heat shock protein family and cytoplasmic proteins by conventional techniques such as phage display library screening. However, little is known about the binding motif based on kinetic parameters determined by surface plasmon resonance analysis. We investigated the major inducible cytosolic 70-kDa heat shock protein (Hsp70)-binding motif with the human leukocyte antigen B*2702-derived peptide Bw4 (RENLRIALRY) by using a Biacore system based on surface plasmon resonance analysis. The K(D) value of Hsp70-Bw4 interaction was 1.8 x 10(-6) m. Analyses with truncated Bw4 variant peptides showed the binding motif of Hsp70 to be seven residues, LRIALRY. To further study the characteristics of this motif, 126 peptides derived from Bw4, each with single amino acid substitution, were synthesized and analyzed for Hsp70 binding affinity. Interestingly, the Hsp70 binding affinity was abrogated when the residues were substituted for by acidic (Asp and Glu) ones at any position. In contrast, if the substitute residue was aromatic (Trp, Tyr, and Phe) or an Arg residue at any position, Hsp70 binding affinity was maintained. Thus, this study presents a new binding motif between Hsp70 and peptides derived from the natural protein human leukocyte antigen B*2702 and may also elucidate some characteristics of the Hsp70 binding characteristic, enhancing our understanding of Hsp70-binding determinants that may influence diverse cellular and physiological processes.
authors.affiliation: Departments of Surgery, South 1 West 17, Chuo-ku, Sapporo 060-8556, Japan; Departments of Pathology, South 1 West 17, Chuo-ku, Sapporo 060-8556, Japan.
authors.forename: Hideki
authors.initials: H
authors.lastname: Maeda
authors.affiliation: Marine Biomedical Institute, Sapporo Medical University School of Medicine, South 1 West 17, Chuo-ku, Sapporo 060-8556, Japan. Electronic address: hsahara@sapmed.ac.jp.
authors.forename: Hiroeki
authors.initials: H
authors.lastname: Sahara
authors.affiliation: Marine Biomedical Institute, Sapporo Medical University School of Medicine, South 1 West 17, Chuo-ku, Sapporo 060-8556, Japan.
authors.forename: Yoko
authors.initials: Y
authors.lastname: Mori
authors.affiliation: Departments of Pathology, South 1 West 17, Chuo-ku, Sapporo 060-8556, Japan.
authors.forename: Toshihiko
authors.initials: T
authors.lastname: Torigo
authors.affiliation: Departments of Pathology, South 1 West 17, Chuo-ku, Sapporo 060-8556, Japan.
authors.forename: Kenjiro
authors.initials: K
authors.lastname: Kamiguchi
authors.affiliation: Department of Bioinformatics Graduate School of Medicine, Chiba University, 1-8-1, Inohana, Chiba 260-8670, Japan.
authors.forename: Yutaka
authors.initials: Y
authors.lastname: Tamura
authors.affiliation: Departments of Pathology, South 1 West 17, Chuo-ku, Sapporo 060-8556, Japan.
authors.forename: Yasuaki
authors.initials: Y
authors.lastname: Tamura
authors.affiliation: Departments of Surgery, South 1 West 17, Chuo-ku, Sapporo 060-8556, Japan.
authors.forename: Kouichi
authors.initials: K
authors.lastname: Hirata
authors.affiliation: Departments of Pathology, South 1 West 17, Chuo-ku, Sapporo 060-8556, Japan.
authors.forename: Noriyuki
authors.initials: N
authors.lastname: Sato
mesh_terms: D000595:Amino Acid Sequence; D001665:Binding Sites; D015235:HLA-B Antigens / Q000378:metabolism*; D018840:HSP70 Heat-Shock Proteins / Q000737:chemistry*; D006801:Humans; D008969:Molecular Sequence Data; D020349:Surface Plasmon Resonance
chemical_list: D015235:HLA-B Antigens; C042388:HLA-Bw4 antigen; D018840:HSP70 Heat-Shock Proteins