The prediction of host–guest binding affinities with computational modelling is still a challenging task. In the 7th statistical assessment of the modeling of proteins and ligands (SAMPL) challenge, a new host named TrimerTrip was synthesized and the thermodynamic parameters of 16 structurally diverse guests binding to the host were characterized. In the TrimerTrip-guest challenge, only structures of the host and the guests are provided, which indicates that the predictions of both the binding poses and the binding affinities are under assessment. In this work, starting from the binding poses obtained from our previous enhanced sampling simulations in the configurational space, we perform extensive alchemical and end-point free energy calculations to calculate the host–guest binding affinities retrospectively. The alchemical predictions with two widely accepted charge schemes (i.e. AM1-BCC and RESP) are in good agreement with the experimental reference, while the end-point estimates perform poorly in reproducing the experimental binding affinities. Aside from the absolute value of the binding affinity, the rank of binding free energies is also crucial in drug design. Surprisingly, the end-point MM/PBSA method seems very powerful in reproducing the experimental rank of binding affinities. Although the length of our simulations is long and the intermediate spacing is dense, the convergence behavior is not very good, which may arise from the flexibility of the host molecule. Enhanced sampling techniques in the configurational space may be required to obtain fully converged sampling. Further, as the length of sampling in alchemical free energy calculations already achieves several hundred ns, performing direct simulations of the binding/unbinding event in the physical space could be more useful and insightful. More details about the binding pathway and mechanism could be obtained in this way. The nonequilibrium method could also be a nice choice if one insists to use the alchemical method, as the intermediate sampling is avoided to some extent.

用计算模型预测主客结合亲和力仍然是一项具有挑战性的任务。在蛋白质和配体建模 (SAMPL) 挑战的第 7 次统计评估中，合成了一种名为 TrimerTrip 的新宿主，并表征了与宿主结合的 16 种结构不同的客体的热力学参数。在 TrimerTrip-guest 挑战中，只提供了宿主和客人的结构，这表明结合姿势和结合亲和力的预测都在评估中。在这项工作中，从我们之前在配置空间中的增强采样模拟中获得的结合位姿开始，我们进行了广泛的炼金术和终点自由能计算，以回顾性地计算主客体结合亲和力。两种广泛接受的电荷方案（即 AM1-BCC 和 RESP）的炼金术预测与实验参考非常一致，而终点估计在再现实验结合亲和力方面表现不佳。除了结合亲和力的绝对值，结合自由能的等级在药物设计中也很重要。令人惊讶的是，终点 MM/PBSA 方法在再现结合亲和力的实验等级方面似乎非常强大。虽然我们的模拟长度较长，中间间距较密，但收敛行为不是很好，这可能是由于主体分子的柔韧性所致。可能需要配置空间中的增强采样技术以获得完全收敛的采样。更远，由于炼金术自由能计算中的采样长度已经达到数百 ns，因此在物理空间中直接模拟结合/解离事件可能会更有用和更有见地。通过这种方式可以获得有关结合途径和机制的更多细节。如果坚持使用炼金法，非平衡法也可能是一个不错的选择，因为在一定程度上避免了中间采样。