Improved Stopped-Flow Time-Resolved Resonance Raman Spectroscopy Device for Studying Enzymatic Reactions
Improved Stopped-Flow Time-Resolved Resonance Raman Spectroscopy Device for Studying Enzymatic Reactions
复制标题
用于研究酶反应的改进的停流时间分辨共振拉曼光谱装置
DOI：
10.1002/jrs.5100
复制
发表时间：
2017
影响因子：
2.5
通讯作者：
and T. Ogura
中科院分区：
文献类型：
--
作者：
S. Yanagisawa;M. S. Deshpande;S. Hirota;T. Nakagawa;and T. Ogura
使用停止流动混合器构建了改进的停止流动共振拉曼光谱装置，其死区时间为3 ms，混合体积为0.1 mL。该装置使用肌红蛋白进行测试，其中铁基氧血红素与过氧化氢溶液混合后，通过时间分辨共振拉曼光谱监测了一种高价血红素的形成反应。用拉曼光谱法得到的铁酰氧血红素生成速率常数与用常规停流吸收光谱法得到的速率常数吻合良好。这些结果证明，该装置普遍适用于酶-底物反应，其时间分辨率明显高于以往报道。版权所有©2017 John Wiley & Sons， Ltd
An improved stopped‐flow resonance Raman spectroscopy device was constructed using a stopped‐flow mixer with a dead time of 3 ms and a mixing volume of 0.1 mL. The device was tested using myoglobin, where the formation reaction of a high‐valent heme species, ferryl‐oxo heme, was monitored by time‐resolved resonance Raman spectroscopy after mixing a ferric myoglobin solution with a hydrogen peroxide solution. The ferryl‐oxo heme formation rate constant obtained by Raman spectroscopy is in good agreement with the rate constant obtained by conventional stopped‐flow absorption spectroscopy for the same reaction under the same conditions. It is proved by these results that the present device is generally applicable to enzyme–substrate reactions with a significantly higher time resolution than previously reported. Copyright © 2017 John Wiley & Sons, Ltd.