Microsecond-Resolved Infrared Spectroscopy on Nonrepetitive Protein Reactions by Applying Caged Compounds and Quantum Cascade Laser Frequency Combs
作者:Mohamad Javad Norahan、Raphael Horvath、Nathalie Woitzik、Pierre Jouy、Florian Eigenmann、Klaus Gerwert、Carsten Kötting
DOI:10.1021/acs.analchem.1c00666
日期:2021.5.4
Infrared spectroscopy is ideally suited for the investigation of protein reactions at the atomic level. Many systems were investigated successfully by applying Fourier transform infrared (FTIR) spectroscopy. While rapid-scan FTIR spectroscopy is limited by time resolution (about 10 ms with 16 cm–1 resolution), step-scan FTIR spectroscopy reaches a time resolution of about 10 ns but is limited to cyclic
红外光谱法非常适合在原子水平上研究蛋白质反应。通过应用傅立叶变换红外(FTIR)光谱技术成功地研究了许多系统。快速扫描FTIR光谱受时间分辨率限制(16 cm –1约10 ms阶跃扫描FTIR光谱达到约10 ns的时间分辨率,但仅限于在相同条件下可以重复数百次的循环反应。因此,只有经历光循环的可光活化蛋白才有可能实现具有高时间分辨率的FTIR。例如由笼状化合物诱导的大量非重复性反应仅限于毫秒时域。现在,双梳量子级联激光器的出现允许在微秒时域内进行快速反应监测。在这里,我们研究了将这种仪器应用于巨大类别的G蛋白的潜力。我们通过将新技术应用于抑制G i的α亚基,比较了通过FTIR和双梳光谱法监测的笼状化合物诱导的反应蛋白和Gα的更大的蛋白质-蛋白质复合物我与G蛋白信号传导(RGS)的其同源调节器。我们以4μs的时间分辨率和与FTIR相当的波长分辨率观察到了良好的数据质量。这比文献中对G蛋白的任