Studies on the structure and function of lysozyme II. The similar activity of urea-treated and cyanate-treated lysozyme

Abstract

Lysozyme (mucopeptide N-acetylmuramylhydrolase, EC 3.2.1.17) which has been treated with 6 M urea has been tested for its enzymatic activity under varying conditions of pH and salt concentration. For the assay the decrease in turbidity of a suspension of lyophilized Micrococcus lysodeikticus was measured. In 6 M urea, a considerable change in activity takes place over a period of at least 16 weeks. During the first 3 weeks an activation occurs that reaches 150% of the activity of the untreated control (pH 7.0, 5 mM salt) and 250% of the activity exhibited by untreated lysozyme at pH 6.3 in 0.1 M phosphate. After 3 weeks the activity decreases under all conditions of assay with the decrease being very sensitive to the salt concentration of the assay medium. At 8 weeks when assayed at pH 8.0, it is completely inactive at 100 mM salt but still 120% of the untreated control in 5 mM salt. When lysozyme is similarly treated with KCNO, changes in lysozyme activity occur that are much like those observed with urea. In 5 mM cyanate the pH-salt-activity profiles change with time in a manner very close to that observed with 6 M urea. It is concluded (1) that carbamylation of free amino groups by cyanate is responsible for the activity changes in urea-treated lysozyme and (2) that assays for the activity of modified lysozymes should be carried out over a wide range of pH and salt concentration.