Barron Christiaan C, Sponagle Brandon J D, Arivalagan Pugazhendhi, D'Cunha Godwin B
Dalhousie University, Halifax, NS, B3H 4J2, Canada.
Department of Chemistry, Cape Breton University, 1250 Grand Lake Road, Sydney, Nova Scotia, B1P 6L2, Canada.
Enzyme Microb Technol. 2017 Jan;96:151-156. doi: 10.1016/j.enzmictec.2016.10.010. Epub 2016 Oct 18.
Phenylalanine ammonia lyase (E.C.4.3.1.24, PAL) activity of Rhodotorula glutinis yeast has been demonstrated in four commonly used ionic liquids. PAL forward reaction was carried out in 1-butyl-3-methylimidazolium methyl sulfate ([BMIM][MeSO]), 1-butyl-3-methylimidazolium tetrafluoroborate ([BMIM][BF]), 1-butyl-3-methylimidazolium hexafluorophosphate ([BMIM][PF]) and 1-butyl-3-methylimidazolium lactate ([BMIM][lactate]). Our experiments have revealed that PAL is catalytically active in ionic liquids and the enzyme activity in ([BMIM][PF]) is comparable to that obtained in aqueous buffer medium. Different conditions were optimized for maximal PAL forward activity including time of incubation (30.0min)-phenylalanine substrate concentration (30.0mM), nature of buffer (50.0mM Tris-HCl), pH (9.0), temperature (37°C), and speed of agitation (100 rev min). Under these optimized conditions, about 83% conversion of substrate to product was obtained for the PAL forward reaction that was determined using UV spectroscopy at 290nm. PAL reverse reaction in ([BMIM][PF]) was determined spectrophotometrically at 520nm; and about 59% substrate conversion was obtained. This data provides further knowledge in enzyme biocatalysis in non-aqueous media, and may be of importance when studying the function of other oligomeric/multimeric proteins and enzymes in ionic liquids.
已在四种常用离子液体中证实了粘红酵母的苯丙氨酸解氨酶（E.C.4.3.1.24，PAL）活性。PAL正向反应在硫酸甲酯1-丁基-3-甲基咪唑鎓（[BMIM][MeSO]）、四氟硼酸1-丁基-3-甲基咪唑鎓（[BMIM][BF]）、六氟磷酸1-丁基-3-甲基咪唑鎓（[BMIM][PF]）和乳酸1-丁基-3-甲基咪唑鎓（[BMIM][乳酸]）中进行。我们的实验表明，PAL在离子液体中具有催化活性，且在（[BMIM][PF]）中的酶活性与在水性缓冲介质中获得的活性相当。针对最大PAL正向活性优化了不同条件，包括孵育时间（30.0分钟）、苯丙氨酸底物浓度（30.0 mM）、缓冲液性质（50.0 mM Tris-HCl）、pH（9.0）、温度（37°C）和搅拌速度（100转/分钟）。在这些优化条件下，使用紫外光谱在290nm处测定，PAL正向反应的底物向产物转化率约为83%。在（[BMIM][PF]）中PAL的逆向反应通过分光光度法在520nm处测定，底物转化率约为59%。这些数据为非水介质中的酶生物催化提供了更多知识，在研究离子液体中其他寡聚/多聚蛋白质和酶的功能时可能具有重要意义。
