Hydrolysis of 4-Methylumbelliferyl Tetra-N-acetyl-β-chitotetraoside by Lysozyme and Its Inhibition by N, N', N"-Triacetylchitotriose

抄録

The hydrolytic activity of lysozyme towards 4-methylumbelliferyl tetra-N-acetyl-β-chitotetraoside (4-MU-(GlcNAc)₄) was little affected by ionic strength, though the activity of lysozyme towards cell suspension of Micrococcus lysodeikticus varied markedly with ionic strength. About 40-60% of lysozyme activity with 4-MU-(GlcNAc)₄ as a substrate was inhibited by 0.1mM N, N', N"-triacetylchitotriose ((GlcNAc)₃), but the lytic activity of lysozyme towards M. lysodeikticus was little affected. The kinetics of hydrolysis of 4-MU-(GlcNAc)₄ by hen egg-white (HEW) lysozyme and human placental (HP) lysozyme and the inhibition of this hydrolysis by (GlcNAc)₃ were investigated. The K₅ values for 4-MU-(GlcNAc)₄ of HEW-and HP-lysozymes were 19.7 and 27.9μM, respectively, and the V_max values were 0.124 and 0.0833 nmol/min/mg, respectively. The k values of both enzymes were very low, implying a poor orientation of the scissile bond in the substrate molecule with respect to the active site of lysozyme. (GlcNAc)₃ inhibited lysozyme with hyperbolic mixed-type inhibition. The inhibition reduced the V_max values of both lysozymes. The K₅ value of HEW-lysozyme was increased by the addition of the inhibitor, whereas the K₅ value of HP-lysozyme was decreased. The K_i value was 29.6μM for HEW-lysozyme and 106μM for HP-lysozyme.