Department of Chemistry and Biochemistry, Baylor University, Waco, Texas 76706, United States.
Department of Chemistry, Trinity University, San Antonio, Texas 78212, United States.
J Am Chem Soc. 2023 Aug 2;145(30):16488-16497. doi: 10.1021/jacs.3c03006. Epub 2023 Jul 24.
The degree to which redox-driven proton pumps regulate net charge during electron transfer (ΔZ) remains undetermined due to difficulties in measuring the net charge of solvated proteins. Values of ΔZ can reflect reorganization energies or redox potentials associated with ET and can be used to distinguish ET from proton(s)-coupled electron transfer (PCET). Here, we synthesized protein "charge ladders" of a Rieske [2Fe-2S] subunit from (truncRp) and made 120 electrostatic measurements of ΔZ across pH. Across pH 5-10, truncRp is suspected of transitioning from ET to PCET, and then to two proton-coupled ET (2PCET). Upon reduction, we found that truncRp became more negative at pH 6.0 by one unit (ΔZ = -1.01 ± 0.14), consistent with single ET; was isoelectric at pH 8.8 (ΔZ = -0.01 ± 0.45), consistent with PCET; and became more positive at pH 10.6 (ΔZ = +1.37 ± 0.60), consistent with 2PCET. These ΔZ values are attributed to protonation of H154 and H134. Across pH, redox potentials of Rp (measured previously) correlated with protonation energies of H154 and H134 and ΔZ for truncRp, supporting a discrete proton pumping mechanism for Rieske proteins at the Fe-coordinating histidines.
由于难以测量溶剂化蛋白质的净电荷，氧化还原驱动的质子泵在电子转移过程中调节净电荷（ΔZ）的程度仍未确定。ΔZ 值可以反映与 ET 相关的重组能或氧化还原电位，可用于区分 ET 与质子（s）耦合电子转移（PCET）。在这里，我们合成了来自 （truncRp）的 Rieske [2Fe-2S] 亚基的蛋白质“电荷梯”，并在 pH 值范围内进行了 120 次静电测量ΔZ。在 pH 5-10 范围内，truncRp 被怀疑从 ET 过渡到 PCET，然后过渡到两个质子耦合 ET（2PCET）。还原后，我们发现 pH 6.0 时 truncRp 变得更负一个单位（ΔZ = -1.01 ± 0.14），与单 ET 一致；在 pH 8.8 时等电（ΔZ = -0.01 ± 0.45），与 PCET 一致；在 pH 10.6 时变得更正（ΔZ = +1.37 ± 0.60），与 2PCET 一致。这些 ΔZ 值归因于 H154 和 H134 的质子化。在整个 pH 值范围内，Rp 的氧化还原电位（以前测量过）与 H154 和 H134 的质子化能以及 truncRp 的 ΔZ 相关，这支持了在 Fe 配位组氨酸处的 Rieske 蛋白具有离散的质子泵浦机制。
