Adsorption of human lysozyme onto hydroxyapatite

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.