Alcantara Juan, Stix Robyn, Huang Katherine, Connor Acadia, East Ray, Jaramillo-Martinez Valeria, Stollar Elliott J, Ball K Aurelia
Department of Chemistry, Skidmore College, Saratoga Springs, NY, United States.
Department of Neuroscience and Pharmacology, Texas Teach University Health Science Center, Lubbock, TX, United States.
Front Mol Biosci. 2021 Nov 15;8:734169. doi: 10.3389/fmolb.2021.734169. eCollection 2021.
Disordered proline-rich motifs are common across the proteomes of many species and are often involved in protein-protein interactions. Proline is a unique amino acid due to the covalent bond between the backbone nitrogen and the proline side chain. The resulting five-membered ring allows proline to sample the state about its peptide bond, which other residues cannot do as readily. Because proline-rich disordered sequences exist as ensembles that likely include structures with the proline peptide bond in , a robust methodology to accurately account for these conformations in the overall ensemble is crucial. Observing the conformations of proline in a disordered sequence is challenging both experimentally and computationally. Nitrogen-hydrogen NMR spectroscopy cannot directly observe proline residues, which lack an amide bond, and computational methods struggle to overcome the large kinetic barrier between the and states, since isomerization usually occurs on the order of seconds. In the current work, Gaussian accelerated molecular dynamics was used to overcome this free energy barrier and simulate proline isomerization in a tetrapeptide (KPTP) and in the 12-residue proline-rich SH3 binding peptide, ArkA. We found that Gaussian accelerated molecular dynamics, when combined with a lowered peptide bond dihedral angle potential energy barrier (15 kcal/mol), allowed sufficient sampling of the proline and states on a microsecond timescale. All ArkA prolines spend a significant fraction of time in , leading to a more compact ensemble with less polyproline II helix structure than an ArkA ensemble with all peptide bonds in . The ensemble containing prolines also matches more closely to circular dichroism data than the all- ensemble. The ability of the ArkA prolines to isomerize likely affects the peptide's ability to bind its partner SH3 domain, and should be studied further. This is the first molecular dynamics simulation study of proline isomerization in a biologically relevant proline-rich sequence that we know of, and a similar protocol could be applied to study multi-proline isomerization in other proline-containing proteins to improve conformational diversity and agreement with data.
富含脯氨酸的无序基序在许多物种的蛋白质组中普遍存在，且常常参与蛋白质-蛋白质相互作用。脯氨酸是一种独特的氨基酸，因为其主链氮与脯氨酸侧链之间存在共价键。由此形成的五元环使脯氨酸能够对其肽键的状态进行采样，而其他残基则无法如此轻易地做到。由于富含脯氨酸的无序序列以可能包含脯氨酸肽键处于 状态的结构的集合形式存在，因此一种能够在整个集合中准确考虑这些构象的稳健方法至关重要。在无序序列中观察脯氨酸的 构象在实验和计算方面都具有挑战性。氮-氢核磁共振光谱无法直接观察到缺乏酰胺键的脯氨酸残基，并且计算方法难以克服 态和 态之间的巨大动力学障碍，因为异构化通常发生在秒级。在当前的工作中，使用高斯加速分子动力学来克服这个自由能障碍，并模拟四肽（KPTP）和富含12个残基的富含脯氨酸的SH3结合肽ArkA中的脯氨酸异构化。我们发现，高斯加速分子动力学与降低的肽键二面角势能障碍（15千卡/摩尔）相结合时，能够在微秒时间尺度上对脯氨酸的 和 态进行充分采样。所有ArkA脯氨酸在 态中花费相当长的时间，导致与所有肽键处于 态的ArkA集合相比，形成了一个更紧凑的集合，且聚脯氨酸II螺旋结构更少。包含 脯氨酸的集合也比全 集合更符合 圆二色性数据。ArkA脯氨酸异构化的能力可能会影响该肽与其伴侣SH3结构域结合的能力，应进一步研究。据我们所知，这是首次对生物学相关的富含脯氨酸序列中的脯氨酸异构化进行分子动力学模拟研究，类似的方案可应用于研究其他含脯氨酸蛋白质中的多脯氨酸异构化，以改善构象多样性并与 数据达成一致。
