Stability of phage T4 lysozymes I. Native properties and thermal stability of wild type and two mutant lysozymes

Abstract

Two mutants of phage T4 lysozyme were prepared and characterized. One mutation substituted a tyrosine residue for tryptophan at position 138. The other substituted tyrosines at all three tryptophan positions of the wild type molecule (126, 138, 158). Comparative studies of the physical properties (absorption, flourescence, circular dichroism) of the three enzymes were performed as a function of pH. Also, the proteins were reversibly melted as a function of pH. Since the unfolding reaction appeared to be a two-state process for all these proteins, the data were analyzed by the van 't Hoff procedure. The changes in stability and activity produced by substitution of Trp 138 were especially significant. The other substitutions were neutral. See the end of the paper for a summary of conclusions. In the appendix the appropriate thermodynamic relations are developed for a constant ΔC P transition. Parts of this material were presented at the meeting of the Society of Biological Chemists in San Francisco in June, 1976 and at the International Conference on Biomolecules at Regensburg, G.F.R. in October, 1976.