Sineshchekov V, Hughes J, Hartmann E, Lamparter T
Biology Department, M. V. Lomonosov Moscow State University, Russia.
Photochem Photobiol. 1998 Feb;67(2):263-7. doi: 10.1562/0031-8655(1998)067<0263:faporp>2.3.co;2.
Fluorescence and photochemical properties of phytochrome from the cyanobacterium Synechocystis were investigated in the temperature interval from 293 to 85 K. The apoprotein was obtained by overexpression in Escherichia coli and assembled to a holophytochrome with phycocyanobilin (PCB) and phytochromobilin (P phi B), Syn(PCB)phy and Syn(P phi B)phy, respectively. Its red-absorbing form, Pr, is characterized at 85 K by the emission and excitation maxima at 682 and 666 nm in Syn(PCB)phy and at 690 and 674 nm in Syn(P phi B)phy. At room temperature, the spectra are blue shifted by 5-10 nm. The fluorescence intensity dropped down by approximately 15-20-fold upon warming from 85 to 293 K and activation energy of the fluorescence decay was estimated to be ca 5.4 and 4.9 kJ mol-1 in Syn(PCB)phy and Syn(P phi B)phy, respectively. Phototransformation of Pr upon red illumination was observed at temperatures above 160-170 K in Syn(PCB)phy and above 140-150 K in Syn(P phi B)phy with a 2-3 nm shift of the emission spectrum to the blue and increase of the intensity of its shorter wavelength part. This was interpreted as a possible formation of the photoproduct of the meta-Ra type of the plant phytochrome. At ambient temperatures, the extent of the Pr phototransformation to the far-red-absorbing form, Pfr, was ca 0.7-0.75 and 0.85-0.9 for Syn(PCB)phy and Syn(P phi B)phy, respectively. Fluorescence of Pfr and of the photoproduct similar to lumi-R was not observed. With respect to the photochemical parameters, Syn(PCB)phy and Syn(P phi B)phy are similar to each other and also to a small fraction of phyA (phyA") and to phyB. The latter were shown to have low photochemical activity at low temperatures in contrast to the major phyA pool (phyA'), which is distinguished by the high extent (ca 50%) of Pr phototransformation at 85 K. These photochemical features are interpreted in terms of different activation barriers for the photoreaction in the Pr excited state.
研究了蓝藻集胞藻中光敏色素在293至85 K温度区间内的荧光和光化学性质。脱辅基蛋白通过在大肠杆菌中过表达获得，并分别与藻蓝胆素（PCB）和藻光敏色素（P phi B）组装成全光敏色素，即Syn(PCB)phy和Syn(P phi B)phy。其红光吸收形式Pr在85 K时，Syn(PCB)phy的发射和激发最大值分别在682和666 nm，Syn(P phi B)phy的发射和激发最大值分别在690和674 nm。在室温下，光谱蓝移5 - 10 nm。从85 K升温至293 K时，荧光强度下降约15 - 20倍，Syn(PCB)phy和Syn(P phi B)phy中荧光衰减的活化能估计分别约为5.4和4.9 kJ mol-1。在Syn(PCB)phy中，温度高于160 - 170 K时，以及在Syn(P phi B)phy中，温度高于140 - 150 K时，观察到Pr在红光照射下发生光转化，发射光谱向蓝光方向移动2 - 3 nm，且其较短波长部分的强度增加。这被解释为可能形成了植物光敏色素的间-Ra型光产物。在环境温度下，Syn(PCB)phy和Syn(P phi B)phy中Pr光转化为远红光吸收形式Pfr的程度分别约为0.7 - 0.75和0.85 - 0.9。未观察到Pfr和类似于lumi-R的光产物的荧光。关于光化学参数，Syn(PCB)phy和Syn(P phi B)phy彼此相似，也与一小部分phyA（phyA"）和phyB相似。与主要的phyA库（phyA'）相比，后者在低温下光化学活性较低，phyA'在85 K时Pr光转化程度较高（约50%）。这些光化学特征是根据Pr激发态光反应的不同活化能垒来解释的。
