化学
傅里叶变换红外光谱
基质(水族馆)
光谱学
氢键
分析化学(期刊)
红外光谱学
黄素组
分子
红外线的
酶
色谱法
生物化学
有机化学
光学
物理
地质学
海洋学
量子力学
作者
Daichi Yamada,Hideki Kandori
出处
期刊:Methods in molecular biology
日期:2014-01-01
卷期号:: 361-376
被引量:14
标识
DOI:10.1007/978-1-4939-0452-5_14
摘要
Light-induced difference Fourier transform infrared (FTIR) spectroscopy is a powerful, sensitive, and informative method to study structure-function relationships in photoreceptive proteins. Strong absorption of water in the IR region is always problematic in this method, but if water content in the sample is controlled during measurements, this method can provide useful information on a single protein-bound water molecule. We established three kinds of sample preparations: hydrated film, redissolved sample, and concentrated solution. Hydrated films were used for the measurements of LOV and BLUF domains, where accurate difference FTIR spectra were obtained in the whole mid-IR region (4,000-800 cm(-1)). Vibrations of S-H stretch of cysteine, O-H stretch of water, and O-H stretch of tyrosine provide important information on hydrogen bonds in these proteins. Redissolved samples were used for the measurements of (6-4) photolyase, in which enzymatic turnover takes place. From the illumination time-dependence of excess amount of substrate, it is possible to isolate the signal originating from the binding of enzyme to substrate. If proteins are less tolerant to drying, as for example cryptochromes of the DASH type, concentrated solution is used. Detailed methodological aspects in light-induced difference FTIR spectroscopy are reviewed by mainly focusing on our results.
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