MEASUREMENT AND GLOBAL ANALYSIS OF THE ABSORBENCY CHANGES IN THE PHOTOCYCLE OF THE PHOTOACTIVE YELLOW PROTEIN FROM ECTOTHIORHODOSPIRA-HALOPHILA
MEASUREMENT AND GLOBAL ANALYSIS OF THE ABSORBENCY CHANGES IN THE PHOTOCYCLE OF THE PHOTOACTIVE YELLOW PROTEIN FROM ECTOTHIORHODOSPIRA-HALOPHILA
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DOI：
10.1016/s0006-3495(94)80643-5
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发表时间：
1994-10-01
影响因子：
3.4
通讯作者：
HELLINGWERF, KJ
中科院分区：
文献类型：
--
作者：
HOFF, WD;VANSTOKKUM, IHM;HELLINGWERF, KJ
采用时间分辨差分吸收光谱法研究了嗜盐外硫红螺菌（Ectothiorhodospira halophila）光活性黄蛋白（PYP）在300-600 nm波长范围内的光循环。测量了时间选通光谱和单波长迹线。数据的整体分析确定，在5 ns和2 s之间的时域中，如之前已经提出的，在PYP的室温光循环中仅涉及两个中间体（Meyer T. E、E. Yakali，M. A. Cusanovich和G.托林。1987.生物化学。26：418-423; Meyer，T. E、G. Tollin，T. P. Causgrove，P. Cheng和R. E.布兰肯希普1991. Biophys. J. 59：988-991）。第一，红移的中间衰减双指数（60%与τ = 0.25毫秒和40%与τ = 1.2毫秒）的蓝移中间。光循环的最后一步是双指数（tau = 0.15 s时为93%，tau = 2.0 s时为7%）恢复到蛋白质的基态。这些光中间体的绝对光谱的重建产生了约465和355 nm的吸收最大值的红色和蓝移的中间体与λ（最大值）在约50%和40%相对于λ（最大值）的基态。这里描述的PYP中的光循环的定量分析铺平了道路，以详细的生物物理分析的过程中发生的感光分子。
The photocycle of the photoactive yellow protein (PYP) from Ectothiorhodospira halophila was examined by time-resolved difference absorption spectroscopy in the wavelength range of 300-600 nm. Both time-gated spectra and single wavelength traces were measured. Global analysis of the data established that in the time domain between 5 ns and 2 s only two intermediates are involved in the room temperature photocycle of PYP, as has been proposed before (Meyer T. E., E. Yakali, M. A. Cusanovich, and G. Tollin. 1987. Biochemistry. 26:418-423; Meyer, T. E., G. Tollin, T. P. Causgrove, P. Cheng, and R. E. Blankenship. 1991. Biophys. J. 59:988-991). The first, red-shifted intermediate decays biexponentially (60% with tau = 0.25 ms and 40% with tau = 1.2 ms) to a blue-shifted intermediate. The last step of the photocycle is the biexponential (93% with tau = 0.15 s and 7% with tau = 2.0 s) recovery to the ground state of the protein. Reconstruction of the absolute spectra of these photointermediates yielded absorbance maxima of about 465 and 355 nm for the red- and blue-shifted intermediate with an epsilon(max) at about 50% and 40% relative to the epsilon(max) of the ground state. The quantitative analysis of the photocycle in PYP described here paves the way to a detailed biophysical analysis of the processes occurring in this photoreceptor molecule.