Tyrosine, cysteine, and proton coupled electron transfer in a ribonucleotide reductase-inspired beta hairpin maquette
Tyrosine, cysteine, and proton coupled electron transfer in a ribonucleotide reductase-inspired beta hairpin maquette
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DOI：
10.1039/c9cc04067f
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发表时间：
2019-08-14
影响因子：
4.9
通讯作者：
Barry, Bridgette A.
中科院分区：
文献类型：
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作者：
McCaslin, Tyler G.;Pagba, Cynthia, V;Barry, Bridgette A.
酪氨酸残基是蛋白质中质子偶联电子转移反应（PCET）的中间体。例如，在核糖核苷酸还原酶（RNR）中，酪氨酸自由基通过含有多个芳香基团的35埃途径氧化活性位点半胱氨酸。单线态酪氨酸被氧化时，自由基变成强酸，可用质子转移反应与氧化还原反应相结合来控制反应速率。在这里，我们描述了一个含有酪氨酸的β发夹，肽O，它的单个酪氨酸Y5和半胱氨酸（C14）之间具有交叉链，非共价相互作用。圆二色性提供了热稳定性β转的证据。EPR光谱分析表明，肽O在160 K紫外光解后形成中性酪氨酸自由基。分子动力学模拟支持酪氨酸单线态和自由基态的酚类/SH相互作用。差分脉冲伏安法显示pH依赖性与中性酪氨酸自由基的形成和其他两个残基的pK(a)变化一致。氧化还原偶联的半胱氨酸pK(A)从9（单线态）降至6.9（自由基）。在pd11下，紫外光解后的皮秒瞬态吸收光谱通过电子转移（ET）监测酪氨酸自由基的重组。肽O的ET速率与14位含有组氨酸和环己基丙氨酸（Cha）的肽的ET速率难以区分。然而，在pD 9，酪氨酸自由基通过PCET衰变，并且与组氨酸14变体相比，衰变速率减慢。值得注意的是，与Cha 14变体相比，衰变速率加快了。我们得出结论，酪氨酸和半胱氨酸之间的氧化还原偶联可以作为蛋白质中PCET的控制机制。
Tyrosine residues act as intermediates in proton coupled electron transfer reactions (PCET) in proteins. For example, in ribonucleotide reductase (RNR), a tyrosyl radical oxidizes an active site cysteine via a 35 angstrom pathway that contains multiple aromatic groups. When singlet tyrosine is oxidized, the radical becomes a strong acid, and proton transfer reactions, which are coupled with the redox reaction, may be used to control reaction rate. Here, we characterize a tyrosine-containing beta hairpin, Peptide O, which has a cross-strand, noncovalent interaction between its single tyrosine, Y5, and a cysteine (C14). Circular dichroism provides evidence for a thermostable beta-turn. EPR spectroscopy shows that Peptide O forms a neutral tyrosyl radical after UV photolysis at 160 K. Molecular dynamics simulations support a phenolic/SH interaction in the tyrosine singlet and radical states. Differential pulse voltammetry exhibits pH dependence consistent with the formation of a neutral tyrosyl radical and a pK(a) change in two other residues. A redox-coupled decrease in cysteine pK(a) from 9 (singlet) to 6.9 (radical) is assigned. At pD 11, picosecond transient absorption spectroscopy after UV photolysis monitors tyrosyl radical recombination via electron transfer (ET). The ET rate in Peptide O is indistinguishable from the ET rates observed in peptides containing a histidine and a cyclohexylalanine (Cha) at position 14. However, at pD 9, the tyrosyl radical decays via PCET, and the decay rate is slowed, when compared to the histidine 14 variant. Notably, the decay rate is accelerated, when compared to the Cha 14 variant. We conclude that redox coupling between tyrosine and cysteine can act as a PCET control mechanism in proteins.