Hen egg-white lysozyme and T4 lysozyme (T4L) contribute to the hydrolysis of peptidoglycans and, recently, have been found to play a vital role in the catalysis of 2-ethynylpyridine (2-EP) polymerization. Both proteins have the ability to produce synthetic polymers derived from a reaction immersed in water; this deviates from the traditional method of polymerization that uses metals and other organic solvents. This project hopes to investigate, by using mutations of T4L, the structure-activity relations required for 2-EP polymerization. Of bacteriophage T4 lysozyme, mutation E11Q provides a unique perspective into the catalytic mechanism of 2-EP polymerization. Unlike WT* and D20A, two variations of the T4 protein investigated within our research group, this mutant lysozyme does not lyse under standard conditions. This remarkable difference led to the development of a protocol to discover how this protein responds in the presence of 2-EP, as well as, how it correlates to the active, binding sites of the amino acid chain. This goal was achieved through the use of CM- and SP-Sepharose purifications, a size-exclusion chromatography, UV/Vis and a fluorescence assay to quantify lytic activity. The results found E11Q to be catalytically inactive for 2-EP polymerization, rendering the protein inept. This significance suggests that the catalytic and lytic activity share a common binding site. The further study of this mutation will utilize HSQC, to determine folding and protein-ligand interactions. Further mutations will allow us to understand a possible mechanism and realize the potential for generating novel materials.

Elimination of 2-ethynylpyridine polymerization using E11Q, a mutation of bacteriophage T4 lysozyme

2019 Spring National Meeting

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