Nano- and microsecond time-resolved FTIR spectroscopy of the halorhodopsin photocycle.
Nano- and microsecond time-resolved FTIR spectroscopy of the halorhodopsin photocycle.
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盐视紫红质光循环的纳秒和微秒时间分辨 FTIR 光谱。
DOI：
10.1111/j.1751-1097.1997.tb03220.x
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发表时间：
1997
影响因子：
3.3
通讯作者：
Braiman,MS
中科院分区：
文献类型：
--
作者：
Dioumaev,AK;Braiman,MS
将时间分辨率为 50 ns 的步进扫描傅立叶变换红外光谱应用于 3-42° C 温度范围内的盐视紫红质 (hR) 光循环的早期阶段。通过全局拟合的动力学数据分析揭示了与早期红移光产物 hK 弛豫相关的两个不同的动力学过程；这些过程在 20°C 时的时间常数 T1⋍ 280 ns 和 T2⋍ 360 μs。光谱特征表明 T1 过程对应于两种不同的 BATHO 中间体 hKE 和 hKL 之间的转变。与 T1 和 T2 跃迁相关的振动差带分布在整个 1800‐900 cm−1 范围内。然而，最大的条带对应于乙烯 C=C 拉伸、指纹 C-C 拉伸和视网膜发色团的氢平面外 (HOOP) 波动。这些差异带的时间演化表明 T1 和 T2 衰变过程主要涉及发色团及其直接环境的松弛。强烈的 HOOP 振动的衰减几乎在 T1 和 T2 过程之间平均分配，表明复杂的生色团松弛，从初级（hKE）bathointermediate 中的扭曲非松弛构象，到 hKL 中较少扭曲的结构，最后到低色位移 hL 中间体中的大致平面结构。 T1 过程中观察到的意外大的激活正熵也支持了这一结论。从 hKE 到 hL 的两次弛豫在很大程度上类似于细菌视紫红质光循环中的相应弛豫 (KE→KL→L)，只是第二步在 hR 中减慢了 200 倍以上。
Step‐scan Fourier transform infrared spectroscopy with 50 ns time resolution was applied to the early stages of the photocycle of halorhodopsin (hR) for the temperature range 3‐42° C. Kinetic data analysis with global fitting revealed two distinct kinetic processes associated with relaxations of the early red‐shifted photoproduct hK; these processes have time constants T1⋍ 280 ns and T2⋍ 360 μs at 20°C. Spectral features demonstrate that the T1process corresponds to a transition between two distinct bathointermediates, hKEand hKL. The vibrational difference bands associated with both T1and T2transitions are spread throughout the whole 1800‐900 cm−1range. However, the largest bands correspond to ethylenic C=C stretches, fingerprint C‐C stretches and hydrogen out‐of‐plane (HOOP) wags of the retinal chromophore. The time evolution of these difference bands indicate that both the T1and T2decay processes involve principally a relaxation of the chromophore and its immediate environment. The decay of the intense HOOP vibrations is nearly equally divided between the T1and T2processes, indicating a complex chromophore relaxation from a twisted nonrelaxed conformation in the primary (hKE) bathointermediate, to a less‐twisted structure in hKL, and finally to a roughly planar structure in the hypsochromically shifted hL intermediate. This conclusion is also supported by the unexpectedly large positive entropy of activation observed for the T1process. The two relaxations from hKEto hL are largely analogous to corresponding relaxations (KE→ KL→ L) in the bacterior‐hodopsin photocycle, except that the second step is slowed down by over 200‐fold in hR.