Binding of N-Acetyl-Chitotriose to Asp 52-Esterified Hen Lysozyme1

Abstract

The pH dependence of the binding constant of (GlcNAc)₃ to Asp 52-esterifled lysozyme was determined by the fluorescence technique. The pK values of Asp 101 in the modified lysozyme and its complex with (GlcNAc)₃ were determined to be 4·5 and 3·6, respectively, at 25°C and 0·1 ionic strength. This result is different from that obtained by Parsons and Raftery ((1972) Biochemistry 11, 1633–1638), who observed no pK shift of Asp 101. The macroscopic pK value of Asp 52 in intact lysozyme determined by them using the pH difference titration data of Asp 52-esterified lysozyme relative to intact lysozyme ((1972) Biochemistry 11, 1623–1629) was 4·5, which is higher by about one pH unit than the pK value determined by our group (Kuramitsu et al. (1974) J. Biochem. 76, 671–683; (1977) ibid. 82, 585–597; (1978) ibid. 83, 159–170). We found that their pH difference titration data in the absence and presence of saccharides can be consistently interpreted in terms of our pK values of Asp 52, Glu 35, and Asp 101, if we assume that the pK value of another ionizable group (probably Asp 48) is perturbed on esterification of Asp 52.