Biosci. Biotech. Biochem. 58 : 174-177 (1994)

FunctionalChanges of Lysozyme by Conjugating with Carboxymethyl Dextran

M. Hattori, S. Imamura, K. Nagasawa and K. Takahashi

Department ofApplied Biological Science,
Faculty of Agriculture,
Tokyo University of Agriculture and Technology
Tokyo 183, Japan

Hen egg lysozyme-carboxymethyl dextran (HEL-CMD) conjugate was prepared by using water-soluble carbodiimide as a model protein-acidic polysaccharide conjugate for improving the protein function. Acid-amide bond between HEL and CMD was confirmed by SDS-PAGE, isoelectric focusing and IR spectra. Molar ratio of CMD to HEL in the conjugate was 1:1. Isoelectric point of the conjugate was 5.5-6.0, which is much lower than that of HEL. Spectroscopic studies suggested that the conformation around Trp residue had not changed but alpha-helix content had decreased to about 1/3 the value for native HEL. The conjugate maintained about 60% of the enzymatic activity of native HEL at 40-60 degree C, while it was about 1.4 times as active as native HEL at 4 degree C and 80 degree C. The conjugate was more stable to proteolysis than native HEL. Denaturation temperature of the conjugate was about 73 degree C, which is almost the same as that of native HEL. However, the enthalpy for denaturation of the conjugate was about 1/3 that of native HEL, which corresponds to the decrease in alpha-helix content.