Perturbation of Short Hydrogen Bonds in Photoactive Yellow Protein via Noncanonical Amino Acid Incorporation
Perturbation of Short Hydrogen Bonds in Photoactive Yellow Protein via Noncanonical Amino Acid Incorporation
复制标题
通过非规范氨基酸掺入扰动光活性黄色蛋白中的短氢键
DOI：
10.1021/acs.jpcb.9b01571
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发表时间：
2019
期刊：
影响因子：
--
通讯作者：
Boxer, Steven G.
中科院分区：
文献类型：
--
作者：
Thomson, Benjamin;Both, Johan;Wu, Yufan;Parrish, Robert M.;Martínez, Todd J.;Boxer, Steven G.
光活性黄蛋白（PYP）是一种小的光感受器蛋白，在去质子化的对香豆酸发色团和两个氨基酸（酪氨酸和谷氨酸）之间有两个异常短的氢键。这导致了关于谷氨酸-发色团氢键是否为低势垒氢键的争论，文献中的结果相互矛盾。我们用琥珀抑制法对酪氨酸的pka2进行了修饰，用化学取代法对发色团进行了修饰。这些修饰蛋白的x射线晶体结构与野生型蛋白几乎相同，因此即使这些修饰改变了供体和受体之间的相对质子亲和力，质子供体和受体之间的重原子距离仍保持不变。尽管质子的相对亲和力发生了相当大的变化，但氢键质子的核磁共振化学位移仅受到适度影响。QM/MM计算用于探索质子的势能面，并将计算的质子位置与经验测量的质子化学位移联系起来。结果与低势垒氢键不一致，但在所有情况下都与局域质子一致，表明离子氢键而不是低势垒氢键。
Photoactive yellow protein (PYP) is a small photoreceptor protein that has two unusually short hydrogen bonds between the deprotonatedp-coumaric acid chromophore and two amino acids, a tyrosine and a glutamic acid. This has led to considerable debate as to whether the glutamic acid-chromophore hydrogen bond is a low barrier hydrogen bond, with conflicting results in the literature. We have modified the pKaof the tyrosine by amber suppression and of the chromophore by chemical substitution. X-ray crystal structures of these modified proteins are nearly identical to the wild-type protein, so the heavy atom distance between proton donor and acceptor is maintained, even though these modifications change the relative proton affinity between donor and acceptor. Despite a considerable change in relative proton affinity, the NMR chemical shifts of the hydrogen-bonded protons are only moderately affected. QM/MM calculations were used to explore the protons’ potential energy surface and connect the calculated proton position with empirically measured proton chemical shifts. The results are inconsistent with a low barrier hydrogen bond but in all cases are consistent with a localized proton, suggesting an ionic hydrogen bond rather than a low barrier hydrogen bond.