title: Adsorption of human lysozyme onto hydroxyapatite. Identification of its adsorbing site using site-directed mutagenesis.
abstract: To elucidate hydroxyapatite-protein interaction, mutant human lysozymes in which the surface charge was modified by site-directed mutagenesis were used. Five mutant human lysozymes (K1A, K13A, K33A, R10A, R14A) were expressed in yeast. The chromatographic behavior of these lysozymes was studied with a HPLC hydroxyapatite column. Elution molarities of K1A and R14A mutants were greatly lowered. While Lys-13 and Arg-10 are located around Lys-1 and Arg-14, K13A and R10A mutants bound onto hydroxyapatite stronger than K1A and R14A mutants. In combination with an X-ray crystal structure of human lysozyme, it is concluded that the adsorbing site of human lysozyme is at the back of the active site and that Arg-14, Lys-1, Arg-10 and Lys-13 play important roles in binding.
authors.affiliation: Division of Biological Sciences, Graduate School of Science, Hokkaido University, Sapporo, Japan.
authors.forename: T
authors.initials: T
authors.lastname: Aizawa
authors.forename: N
authors.initials: N
authors.lastname: Koganesawa
authors.forename: A
authors.initials: A
authors.lastname: Kamakura
authors.forename: K
authors.initials: K
authors.lastname: Masaki
authors.forename: A
authors.initials: A
authors.lastname: Matsuura
authors.forename: H
authors.initials: H
authors.lastname: Nagadome
authors.forename: Y
authors.initials: Y
authors.lastname: Terada
authors.forename: K
authors.initials: K
authors.lastname: Kawano
authors.forename: K
authors.initials: K
authors.lastname: Nitta
mesh_terms: D000327:Adsorption; D000595:Amino Acid Sequence; D019943:Amino Acid Substitution; D001665:Binding Sites; D003001:Cloning, Molecular; D018360:Crystallography, X-Ray; D017886:Durapatite; D006801:Humans; D008958:Models, Molecular; D009113:Muramidase / Q000737:chemistry* / Q000378:metabolism*; D016297:Mutagenesis, Site-Directed; D011487:Protein Conformation; D011994:Recombinant Proteins / Q000737:chemistry / Q000378:metabolism; D012441:Saccharomyces cerevisiae
chemical_list: D011994:Recombinant Proteins; D017886:Durapatite; D009113:Muramidase