19F NMR studies on 8-fluoroflavins and 8-fluoro flavoproteins.
19F NMR studies on 8-fluoroflavins and 8-fluoro flavoproteins.
复制标题
8-氟黄素和 8-氟黄素蛋白的 19F NMR 研究。
DOI：
10.1021/bi00463a008
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发表时间：
1990
期刊：
影响因子：
2.9
通讯作者：
Massey,V
中科院分区：
文献类型：
--
作者：
Macheroux,P;Kojiro,CL;Schopfer,LM;Chakraborty,S;Massey,V
密歇根大学医学院生物化学系，邮政信箱0606，密歇根州安阿伯，邮编48109-0606，接收日期：1989年8月7日;修订后的手稿，接收日期：1989年10月18日摘要：氧化和还原形式的8-氟核黄素、8-氟-FAD和8-氟黄素重构的黄素蛋白黄素氧还蛋白、核黄素结合蛋白、测定D-氨基酸氧化酶、对羟基苯甲酸羟化酶、老黄酶、邻氨基苯甲酸羟化酶、总酰基辅酶A脱氢酶、葡萄糖氧化酶和L-乳酸氧化酶。对于所研究的蛋白质，氧化共振出现在10.1 ppm范围内，而还原共振分布在10.3 ppm范围内。还原引起了约27 ppm的高场位移的游离8-氟代甜菜碱和大多数的8-氟黄素蛋白。值得注意的例外是8-氟-FMN黄素氧还蛋白，其位移为37.6 ppm，表明苯环中的电子密度异常高。配体结合到氧化的8-氟黄素蛋白引起1.5-5 ppm的高场或低场位移，这取决于蛋白质/配体组合。8-氟-FAD邻氨基苯甲酸羟化酶共振向低场移动，并在邻氨基苯甲酸存在下分裂成两个峰。8-氟-FMN老黄酶共振在与形成电荷转移的对位取代酚盐络合后向高场移动。随着酚盐的给电子能力的增加，高场位移从< 1 ppm增加到5 ppm。天然老黄酶与2，4-二氟苯酚络合后，配体的氟共振发生位移，分裂成两对信号。每对信号与不同的老黄酶同工酶相关。
Department of Biological Chemistry, University of Michigan Medical School, PO Box 0606, Ann Arbor, Michigan 48109-0606 Received August 7, 1989; Revised Manuscript Received October 18, 1989 abstract: The 19F NMR spectra of the oxidized and reduced forms of 8-fluororiboflavin, 8-fluoro-FAD, and the 8-fluoroflavin-reconstituted flavoproteins flavodoxin, riboflavin binding protein, D-amino acid oxidase, p-hydroxybenzoate hydroxylase, Old Yellow Enzyme, anthranilate hydroxylase, general acyl-CoA de-hydrogenase, glucose oxidase, and L-lactate oxidase were measured. For the proteins studied the oxidized resonances appeared over a 10.1-ppm range, while the reduced resonances were spread over 10.3 ppm. Reduction caused an upfield shift of about 27 ppm for the free 8-fluoroflavins and most of the 8-fluoro flavoproteins. The notable exception was 8-fluoro-FMN flavodoxin, which was shifted 37.6 ppm, indicating an unusually high electron density in the benzene ring. Ligand binding to the oxidized 8-fluoro flavoproteins caused either upfield or downfield shifts of 1.5-5 ppm, depending on the protein/ligand combination. The 8-fluoro-FAD anthranilate hydroxylase resonance was shifted downfield and split into two peaks in the presence of anthranilate. The 8-fluoro-FMN Old Yellow Enzyme resonance was shifted upfield upon complexation with charge-transfer-forming, para-substituted phenolates. The upfield shift increased from< 1 to 5 ppm as the electron-donating capacity of the phenolate increased. Complexation of native Old Yellow Enzyme with 2, 4-difluorophenol caused the fluorine resonances of the ligand to shift and split into twopairs of signals. Each pair of signals was associated with a different isozyme of Old Yellow Enzyme.