Structural changes caused by site-directed mutagenesis of tyrosine-98 in Desulfovibrio vulgaris flavodoxin delineated by 1H and 15N NMR spectroscopy: implications for redox potential modulation.
Structural changes caused by site-directed mutagenesis of tyrosine-98 in Desulfovibrio vulgaris flavodoxin delineated by 1H and 15N NMR spectroscopy: implications for redox potential modulation.
复制标题
通过 1H 和 15N NMR 光谱描述的 Desulfovibrio vulgaris 黄素氧还蛋白中酪氨酸 98 的定点诱变引起的结构变化：对氧化还原电位调节的影响。
DOI：
10.1021/bi00255a011
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发表时间：
1994
期刊：
影响因子：
2.9
通讯作者：
Swenson,RP
中科院分区：
文献类型：
--
作者：
Stockman,BJ;Richardson,TE;Swenson,RP
摘要：黄曲霉毒素在低氧化还原电势下调节其他蛋白质的修复基之间的电子转移。蛋白质与黄素单核苷酸辅因子的相互作用使氧化/半对苯二酚和半对苯二酚/对苯二酚氧化还原电位明显偏离其自由溶液的值。为了研究98位酪氨酸在这一过程中的可能作用，我们利用异核三维核磁共振谱测定了普通黄毒弧菌的野生型和4个98位突变株Y98W、Y98H、Y98A和Y98R的溶液构象。指定的JH和15N共振表明，蛋白质的二级结构和拓扑结构是相同的。然而，发生大量突变引起的化学位移变化的残基分布在黄素辅因子结合位点上。距离和二面角约束被用来生成野生型和突变型蛋白质的溶液结构。总的来说，突变的蛋白质在黄素结合部位没有明显的构象变化。确实发生的变化很小，是由于适应位置-98的新侧链所需的不同填充相互作用造成的。比较了突变蛋白中黄素结合部位的溶剂可及性和静电性质与野生型结构的溶剂可及性和静电性质。结构数据支持这样的假设，即野生型蛋白质中半喹/对苯二酚对的很低中点在很大程度上是由黄素结合位点非极化环境中黄素氢醌阴离子的能量不利形成所调节的。
Revised Manuscript Received October 16, 1994® abstract: Flavodoxins mediate electron transfer at low redox potential between the prosthetic groups of other proteins. Interactions between the protein and the flavin mononucleotide cofactor shift both the oxidized/semiquinone and semiquinone/hydroquinone redox potentials significantly from their free-insolution values. In order to investigate the possible role thatthe tyrosine at position 98 plays in this process, we have used heteronuclear three-dimensional NMR spectroscopy to determine the solution conformation of wild-type and four position-98 mutants, Y98W, Y98H, Y98A, and Y98R, of Desulfovibrio vulgaris flavodoxin. Assigned JH and 15N resonances indicate that the secondarystructure and topology of the proteins are identical. However, residues that undergo substantial mutation-induced changes in chemical shift are spread throughout the flavin cofactor binding site. Distance and dihedral angle constraints were used to generate solution structures for the wild-type and mutant proteins. Collectively, the mutant proteins have no gross conformational changes in the flavin binding site. The changes that do occur are minor and result from the different packing interactions required to accommodate the new side chain at position-98. The solvent accessibility and electrostatic nature of the flavin binding site in the mutant proteins are compared to those of the wild-type structure. The structural data support the hypothesis that the very low midpoint of the semiquinone/hydroquinone couple in the wild-type protein is modulated to a large extent by the energetically unfavorableformation of the flavin hydroquinone anion in the apolar environment of the flavin binding site.