Importance of van der Waals contact between Glu 35 and Trp 109 to the catalytic action of human lysozyme.

The importance of van der Waals contact between Glu 35 and Trp 109 to the active-site structure and the catalytic properties of human lysozyme (HL) has been investigated by site-directed mutagenesis. The X-ray analysis of mutant HLs revealed that both the replacement of Glu 35 by Asp or Ala, and the replacement of Trp 109 by Phe or Ala resulted in a significant but localized change in the active-site cleft geometry. A prominent movement of the backbone structure was detected in the region of residues 110 to 120 and in the region of residues 100 to 115 for the mutations concerning Glu 35 and Trp 109, respectively. Accompanied by the displacement of the main-chain atoms with a maximal deviation of C alpha atom position ranging from 0.7 A to 1.0 A, the mutant HLs showed a remarkable change in the catalytic properties against Micrococcus luteus cell substrate as compared with native HL. Although the replacement of Glu 35 by Ala completely abolished the lytic activity, HL-Asp 35 mutant retained a weak but a certain lytic activity, showing the possible involvement of the side-chain carboxylate group of Asp 35 in the catalytic action. The kinetic consequence derived from the replacement of Trp 109 by Phe or Ala together with the result of the structural change suggested that the structural detail of the cleft lobe composed of the residues 100 to 115 centered at Ala 108 was responsible for the turnover in the reaction of HL against the bacterial cell wall substrate. The results revealed that the van der Waals contact between Glu 35 and Trp 109 was an essential determinant in the catalytic action of HL.