Tyr30 of amicyanin is not critical for electron transfer to cytochrome c-551i:: implications for predicting electron transfer pathways
Tyr30 of amicyanin is not critical for electron transfer to cytochrome c-551i:: implications for predicting electron transfer pathways
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DOI：
10.1016/s0005-2728(00)00052-9
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发表时间：
2000-02-24
影响因子：
4.3
通讯作者：
Zhu, ZY
中科院分区：
文献类型：
--
作者：
Davidson, VL;Jones, LH;Zhu, ZY
对甲胺脱氢酶-氨蓝蛋白-细胞色素c-551I蛋白电子转移(ET)复合体的通路分析表明，从氨蓝蛋白的I型铜到细胞色素c-551I的血红素，有两组效率相当的ET通路。在一种途径中，电子通过Cys(92)铜配体离开铜，在另一种途径中，它通过Met(98)铜配体离开。如果路径算法被修改为包括来自金属-配体耦合的各向异性的贡献，而与铜-配体键长的差异无关，则通过Cys(92)的路径被预测为比通过任何其他铜配体的路径强至少100倍。所有通过Cys(92)的优选路径都包括从Cys(92)到Tyr(30)侧链的穿过空间跳跃。为了确定通过Cys(92)的途径是否优先用于ET，通过定点突变将Tyr(30)改变为其他氨基酸残基，一些突变的蛋白质非常不稳定，提示Tyr(30)具有稳定蛋白质结构的作用。可以分离和分析Y30F和Y30I突变株的氨基花青素。对于Y30I突变体，倾向于ET Via Cys(92)的改良路径分析预测ET率至少下降两个数量级，而标准Path分析预测ET率没有变化，因为ET Via Met(98)不受影响。在实验上，Y30I和Y30F突变体的ET率与野生型氨基蓝蛋白没有区别，对这些观察到的可能的解释进行了讨论，以及它们对预测金属蛋白ET反应途径的影响，(C)2000 Elsevier Science B.V.，保留所有权利。
A Pathways analysis of the methylamine dehydrogenase-amicyanin-cytochrome c-551i protein electron transfer (ET) complex predicts two sets of ET pathways of comparable efficiency from the type I copper of amicyanin to the heme of cytochrome c-551i. In one pathway, the electron exits copper via the Cys(92) copper ligand, and in the other, it exits via the Met(98) copper ligand. If the Pathways algorithm is modified to include contributions from the anisotropy of metal-ligand coupling, independent of differences in copper-ligand bond length, then the pathways via Cys(92) are predicted to be at least 100-fold more strongly coupled than the pathways via any of the other copper ligands. All of the favored pathways via Cys(92) include a through-space jump from Cys(92) to the side chain of Tyr(30). To determine whether or not the pathways via Cys(92) are preferentially used for ET, Tyr(30) was changed to other amino acid residues by site-directed mutagenesis, Some mutant proteins were very unstable suggesting a role for Tyr(30) in stabilizing the protein structure. Y30F and Y30I mutant amicyanins could be isolated and analyzed. For the Y30I mutant, the modified Pathways analysis which favors ET via Cys(92) predicts a decrease in ET rate of at least two orders of magnitude, whereas the standard Pathways analysis predicts no change in ET rate since ET via Met(98) is not affected. Experimentally, the ET rates of the Y30I and Y30F mutants were indistinguishable from that of wild-type amicyanin, Likely explanations for these observations are discussed as are their implications for predicting pathways for ET reactions of metalloproteins, (C) 2000 Elsevier Science B.V, All rights reserved.