Analysis of flash-induced FTIR difference spectra of the S-state cycle in the photosynthetic water-oxidizing complex by uniform 15N and 13C isotope labeling
Analysis of flash-induced FTIR difference spectra of the S-state cycle in the photosynthetic water-oxidizing complex by uniform 15N and 13C isotope labeling
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DOI：
10.1021/bi0341612
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发表时间：
2003-05-27
期刊：
影响因子：
2.9
通讯作者：
Sugiura, M
中科院分区：
文献类型：
--
作者：
Noguchi, T;Sugiura, M
利用光系统II(PS II)的均匀N-15和C-13同位素标记，分析了光合作用水氧化S态周期的闪光诱导傅里叶变换红外差谱中的蛋白质带。用未标记、N-15标记和C-13标记的细长热核球菌PSⅡ核心复合体的水合(1800-1200 cm(-1)区)或氢化(3500-3100 cm(-1)区)膜获得了第一至第四次闪光照射时的差异光谱。N-15和C-13标记的谱带频率的移动提供了3450-3250和1700-1630 cm(-1)区域的主峰分别归属于多肽骨架的NH伸缩和酰胺I模式，以及1600-1500 cm(-1)区域的一些峰归属于骨架的酰胺II模式。后一区域的其他显著峰和1450-1300 cm(-1)区域的大多数峰在C-13标记时表现出较大的下移，但在N-15标记时没有变化，因此分别归属于Glu、Asp或C-末端的羧酸基的不对称和对称的COO伸缩振动。在第一和第四闪光光谱中，峰的位置彼此对应得很好，第一和/或第二闪光光谱中的大多数谱带在第三和/或第四闪光光谱中出现了相反的强度符号。这一观察表明，S-1-->；S-2和/或S-2-->；S-3转变中的蛋白质运动大多在S-3-->；S-0和/或S-0-S-1转变中发生逆转，这表明水氧化复合体的蛋白质部分起到了催化作用。在S状态循环中，羧酸基结构的剧烈变化表明天冬氨酸和/或谷氨酸侧链在光合作用水氧化反应机制中起着重要作用。
Protein bands in flash-induced Fourier transform infrared (FTIR) difference spectra of the S-state cycle of photosynthetic water oxidation were analyzed by uniform N-15 and C-13 isotopic labeling of photosystem II (PS II). The difference spectra upon first- to fourth-flash illumination were obtained with hydrated (for the 1800-1200 cm(-1) region) or deuterated (for the 3500-3100 cm(-1) region) films of unlabeled, N-15-labeled, and C-13-labeled PS II core complexes from Thermosynechococcus elongatus. Shifts of band frequencies upon N-15 and C-13 labeling provided the assignments of major peaks in the regions of 3450-3250 and 1700-1630 cm(-1) to the NH stretches and amide I modes of polypeptide backbones, respectively, and the assignments of some of the peaks in the 1600-1500 cm(-1) region to the amide II modes of backbones. Other prominent peaks in the latter region and most of the peaks in the 1450-1300 cm(-1) region exhibited large downshifts upon C-13 labeling but were unchanged by N-15 labeling, and hence assigned to the asymmetric and symmetric COO- stretching vibrations, respectively, of carboxylate groups in Glu, Asp, or the C-terminus. Peak positions corresponded well with each other among the first- to fourth-flash spectra, and most of the bands in the first- and/or second-flash spectra appeared with opposite signs of intensity in the third- and/or fourth-flash spectra. This observation indicates that the protein movements in the S-1-->S-2 and/or S-2-->S-3 transitions are mostly reversed in the S-3-->S-0 and/or S-0-S-1 transitions, representing a catalytic role of the protein moieties of the water-oxidizing complex. Drastic structural changes in carboxylate groups over the S-state cycle suggest that the Asp and/or Glu side chains play important roles in the reaction mechanism of photosynthetic water oxidation.