Rothschild K J, Roepe P, Lugtenburg J, Pardoen J A
Biochemistry. 1984 Dec 4;23(25):6103-9. doi: 10.1021/bi00320a031.
The first step of the bacteriorhodopsin (bR) photocycle involves the formation of a red-shifted product, K. Fourier transform infrared difference spectra of the bR570 to K630 transition at 81 K has been measured for bR containing different isotopic substitutions at the retinal Schiff base. In the case of bacteriorhodopsin containing a deuterium substitution at the Schiff base nitrogen, carbon 15, or both, we find spectral changes in the 1600-1610- and 1570-1580-cm-1 region consistent with the hypothesis that the K630 C=N stretching mode of a protonated Schiff base is located near 1609 cm-1. A similar set of Schiff base deuterium substitutions for retinal containing a 13C at the carbon 10 position strongly supports this conclusion. This assignment of the K630 C=N stretching vibration provides evidence that the bR Schiff base proton undergoes a substantial environmental change most likely due to separation from a counterion. In addition, a correlation is found between the C=N stretching frequency and the maximum wavelength of visible absorption, suggesting that movement of a counterion relative to the Schiff base proton is the main source of absorption changes in the early stages of the photocycle. Such a movement is a key prediction of several models of proton transport and energy transduction. Evidence is also presented that one or more COOH groups are involved in the formation of the K intermediate.
细菌视紫红质（bR）光循环的第一步涉及形成一个红移产物K。已测量了在81K下bR570到K630跃迁的傅里叶变换红外差谱，该bR在视黄醛席夫碱处含有不同的同位素取代。对于在席夫碱氮、碳15或两者处含有氘取代的细菌视紫红质，我们发现在1600 - 1610和1570 - 1580 cm⁻¹区域的光谱变化与质子化席夫碱的K630 C=N伸缩模式位于1609 cm⁻¹附近的假设一致。对于在碳10位置含有¹³C的视黄醛进行的一组类似的席夫碱氘取代有力地支持了这一结论。K630 C=N伸缩振动的这种归属提供了证据，表明bR席夫碱质子经历了显著的环境变化，很可能是由于与抗衡离子分离所致。此外，发现C=N伸缩频率与可见吸收的最大波长之间存在相关性，这表明抗衡离子相对于席夫碱质子的移动是光循环早期吸收变化的主要来源。这种移动是几种质子传输和能量转导模型的关键预测。还提供了证据表明一个或多个COOH基团参与了K中间体的形成。
