Recovery of enzyme structure and activity following rehydration from ionic liquid

溶菌酶 离子液体 圆二色性 化学 活动站点 离子 分子动力学 结晶学 生物化学 催化作用 计算化学 有机化学
作者
Pei‐Yin Lee,Onkar Singh,Harry Bermudez,Silvina Matysiak
出处
期刊:Physical Chemistry Chemical Physics [Royal Society of Chemistry]
卷期号:24 (17): 10365-10372 被引量:2
标识
DOI:10.1039/d2cp00608a
摘要

Long-term preservation of proteins at room temperature continues to be a major challenge. Towards using ionic liquids (ILs) to address this challenge, here we present a combination of experiments and simulations to investigate changes in lysozyme upon rehydration from IL mixtures using two imidazolium-based ILs (1-ethyl-3-methylimidazolium ethylsulfate, [EMIM][EtSO4] and 1-ethyl-3-methylimidazolium diethylphosphate, [EMIM][Et2PO4]). Various spectroscopic experiments and molecular dynamics simulations are performed to ascertain the structure and activity of lysozyme. Circular dichroism spectroscopy confirms that lysozyme maintains its secondary structure upon rehydration, even after 295 days. Increasing the IL concentration decreases the activity of lysozyme and is ultimately quenched at sufficiently high IL concentrations, but the rehydration of lysozyme from high IL concentrations completely restores its activity. Such rehydration occurs in the most common lysozyme activity assay, but without careful attention, this effect on the IL concentration can be overlooked. From simulations we observe occupation of [EMIM+] ions near the vicinity of the active site and the ligand-lysozyme complex is less stable in the presence of ILs, which results in the reduction of lysozyme activity. Upon rehydration, fast leaving of [EMIM+] is observed and the availability of active site is restored. In addition, suppression of structural fluctuations is also observed when in high IL concentrations, which also explains the decrease of activity. This structure suppression is recovered after undergoing rehydration. The return of native protein structure and activity indicates that after rehydration lysozyme returns to its original state. Our results also suggest a simple route to protein recovery following extended storage.
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