Abstract：
With recent large-scale applications and validations, the relative binding free energy (RBFE) calculated using alchemical free energy methods has been proven to be an accurate measure to probe the binding of small-molecule drug candidates. On the other hand, given the flexibility of peptides, it is of great interest to find out whether sufficient sampling could be achieved within the typical time scale of such calculation, and a similar level of accuracy could be reached for peptide drugs. However, the systematic evaluation of such calculations on protein-peptide systems has been less reported. Most reported studies of peptides were restricted to a limited number of data points or lacking experimental support. To demonstrate the applicability of the alchemical free energy method for protein-peptide systems in a typical real-world drug discovery project, we report an application of the thermodynamic integration (TI) method to the RBFE calculation of ghrelin receptor and its peptide agonists. Along with the calculation, the synthesis and in vitro EC50 activity of relamorelin and 17 new peptide derivatives were also reported. A cost-effective criterion to determine the data collection time was proposed for peptides in the TI simulation. The average of three TI repeats yielded a mean absolute error of 0.98 kcal/mol and Pearson\'s correlation coefficient (R) of 0.77 against the experimental free energy derived from the in vitro EC50 activity, showing good repeatability of the proposed method and a slightly better agreement than the results obtained from the arbitrary time frames up to 20 ns. Although it is limited by having one target and a deduced binding pose, we hope that this study can add some insights into alchemical free energy calculation of protein-peptide systems, providing theoretical assistance to the development of peptide drugs.
摘要：
随着最近的大规模应用和验证，使用炼金术自由能方法计算的相对结合自由能(RBFE)已被证明是探测小分子候选药物结合的准确量度。另一方面,鉴于肽的灵活性，非常感兴趣的是，找出在这种计算的典型时间尺度内是否可以实现足够的采样，肽类药物也能达到相似的精确度。然而,对蛋白质-肽系统进行此类计算的系统评估报道较少。大多数报道的肽研究仅限于有限数量的数据点或缺乏实验支持。为了证明炼金术自由能方法在典型的现实世界药物发现项目中对蛋白质-肽系统的适用性，我们报道了热力学积分（TI）方法在生长素释放肽受体及其肽激动剂的RBFE计算中的应用。随着计算，还报道了relamorelin和17种新肽衍生物的合成和体外EC50活性。对于TI模拟中的肽，提出了确定数据收集时间的经济有效标准。三个TI重复的平均值产生0.98kcal/mol的平均绝对误差和0.77的皮尔逊相关系数（R）与来自体外EC50活性的实验自由能，显示了所提出方法的良好可重复性，并且比从高达20ns的任意时间范围获得的结果具有更好的一致性。尽管它受到一个靶标和推断的结合姿势的限制，我们希望这项研究可以为蛋白质-肽系统的炼金术自由能计算增加一些见解，为多肽药物的开发提供理论上的帮助。
