Single-Shot Sub-microsecond Mid-infrared Spectroscopy on Protein Reactions with Quantum Cascade Laser Frequency Combs
Single-Shot Sub-microsecond Mid-infrared Spectroscopy on Protein Reactions with Quantum Cascade Laser Frequency Combs
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DOI：
10.1021/acs.analchem.8b02531
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发表时间：
2018-09-04
影响因子：
7.4
通讯作者：
Kottke, Tilman
中科院分区：
文献类型：
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作者：
Klocke, Jessica L.;Mangold, Markus;Kottke, Tilman
不可逆蛋白质反应的动力学分析需要一种分析技术，可以在一次拍摄中获得随时间变化的红外光谱。本文利用量子级联激光器产生的中红外频梳，设计了一种基于双频梳光谱学的光谱仪。分子振动共振对梳子强度的衰减导致指纹区吸收光谱覆盖55 cm（-1）。该装置的原始分辨率为0.3 cm(-1)，噪声水平在μ OD范围内，并实现亚微秒时间分辨率。我们展示了光激活质子泵细菌视紫红质的光谱和瞬态的同时记录。更重要的是，单次拍摄，即单次可见光激发，就足以提取细菌视紫红质光循环中几种中间体的光谱和动力学特征。这一发展为酶转化的无创分析铺平了道路，具有高时间分辨率，广谱覆盖和最小的样品消耗。
The kinetic analysis of irreversible protein reactions requires an analytical technique that provides access to time-dependent infrared spectra in a single shot. Here, we present a spectrometer based on dual-frequency-comb spectroscopy using mid-infrared frequency combs generated by quantum cascade lasers. Attenuation of the intensity of the combs by molecular vibrational resonances results in absorption spectra covering 55 cm(-1) in the fingerprint region. The setup has a native resolution of 0.3 cm(-1), noise levels in the mu OD range, and achieves sub-microsecond time resolution. We demonstrate the simultaneous recording of both spectra and transients of the photoactivated proton pump bacteriorhodopsin. More importantly, a single shot, i.e., a single visible light excitation, is sufficient to extract spectral and kinetic characteristics of several intermediates in the bacteriorhodopsin photocycle. This development paves the way for the noninvasive analysis of enzymatic conversions with high time resolution, broad spectral coverage, and minimal sample consumption.