data for "An unbound proline-rich signaling peptide frequently samples cis conformations in Gaussian accelerated molecular dynamics simulations"

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 <em>cis</em> 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 <em>cis</em>, a robust methodology to accurately account for these conformations in the overall ensemble is crucial. Observing the <em>cis </em>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 <em>cis </em>and <em>trans </em>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 <em>cis </em>and <em>trans </em>states on a microsecond timescale. All ArkA prolines spend a significant fraction of time in <em>cis</em>, leading to a more compact ensemble with less polyproline II helix structure than an ArkA ensemble with all peptide bonds in <em>trans</em>. The ensemble containing <em>cis</em> prolines also matches more closely to <em>in vitro</em> circular dichroism data than the all-<em>trans</em> 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 <em>in vitro</em> data.富含脯氨酸的无序基序广泛存在于众多物种的蛋白质组中，且常参与蛋白质-蛋白质相互作用过程。脯氨酸是一种独特的氨基酸，其骨架氮原子与侧链之间存在共价键，由此形成的五元环使得脯氨酸能够在其肽键处呈现顺式（cis）构象，而其他氨基酸残基则难以实现这一点。由于富含脯氨酸的无序序列以构象系综形式存在，且其中大概率包含脯氨酸肽键为顺式的结构，因此开发一种可在整体系综中准确表征此类构象的稳健方法至关重要。 在无序序列中观测脯氨酸的顺式构象，无论在实验还是计算层面都极具挑战：氮氢核磁共振（NMR）光谱无法直接观测脯氨酸残基，因为后者缺乏酰胺键；而计算方法则难以克服顺式与反式（trans）构象间的巨大动力学能垒，因为脯氨酸的异构化通常仅在秒级时间尺度上发生。 本研究采用高斯加速分子动力学（Gaussian accelerated molecular dynamics）方法克服该自由能能垒，分别在四肽（KPTP）以及含12个残基的富含脯氨酸的SH3结构域结合肽ArkA中模拟脯氨酸的异构化过程。研究发现，当高斯加速分子动力学与降低后的肽键二面角势能垒（15 kcal/mol）相结合时，能够在微秒级时间尺度上对脯氨酸的顺式与反式构象进行充分采样。 ArkA中的所有脯氨酸残基均会有相当比例的时间处于顺式构象，由此形成的构象系综相较于所有肽键均为反式的ArkA系综更为紧凑，且所含的聚脯氨酸II型螺旋结构更少。包含顺式脯氨酸的构象系综，相较于全反式系综，与体外（in vitro）圆二色性（circular dichroism）实验数据的吻合度更高。ArkA的脯氨酸残基发生异构化的能力，可能会影响该肽与其结合伴侣SH3结构域的相互作用，这一问题有待进一步研究。 据我们所知，本研究是首个针对生物学相关的富含脯氨酸序列开展脯氨酸异构化分子动力学模拟的研究，类似的研究方案可被用于其他含脯氨酸蛋白质中的多脯氨酸异构化研究，以提升构象多样性并改善与体外实验数据的吻合度。