Molecular Modeling and Biopharmaceutical Center, College of Pharmacy, University of Kentucky, 789 South Limestone Street, Lexington, Kentucky 40536, United States.
Department of Pharmaceutical Sciences, College of Pharmacy, University of Kentucky, 789 South Limestone Street, Lexington, Kentucky 40536, United States.
J Phys Chem B. 2023 Dec 21;127(50):10723-10729. doi: 10.1021/acs.jpcb.3c06392. Epub 2023 Dec 8.
A combined computational and experimental study has been carried out to explore and test a quantitative correlation relationship between the relative catalytic efficiency (RCE) of human butyrylcholinesrase (BChE) mutant-catalyzed hydrolysis of substrate (-)-cocaine and the total hydrogen bonding energy (tHBE) of the carbonyl oxygen of the substrate with the oxyanion hole of the enzyme in the modeled transition-state structure (TS1), demonstrating a satisfactory linear correlation relationship between ln(RCE) and tHBE. The satisfactory correlation relationship has led us to computationally predict and experimentally confirm new human BChE mutants that have a further improved catalytic activity against (-)-cocaine, including the most active one (the A199S/F227S/S287G/A328W/Y332G mutant) with a 2790-fold improved catalytic efficiency (/ = 2.5 × 10 min M) compared to the wild-type human BChE. Compared to the reference mutant (the A199S/S287G/A328W/Y332G mutant) tested in the reported clinical development of an enzyme therapy for cocaine dependence treatment, this new mutant (with a newly predicted additional F227S mutation) has an improved catalytic efficiency against (-)-cocaine by ∼2.6-fold. The good agreement between the computational and experimental ln(RCE) values suggests that the obtained correlation relationship is robust for computational prediction. A similar correlation relationship could also be explored in studying BChE or other serine hydrolases/esterases with an oxyanion hole stabilizing the carbonyl oxygen in the rate-determining reaction step of the enzymatic hydrolysis of other substrates.
一项结合计算和实验的研究已经进行，以探索和测试人丁酰胆碱酯酶（BChE）突变体催化底物（-）可卡因水解的相对催化效率（RCE）与羰基氧与酶模型过渡态结构（TS1）中的氧阴离子穴之间的总氢键能（tHBE）之间的定量相关关系，证明ln（RCE）和tHBE 之间存在令人满意的线性相关关系。令人满意的相关性使我们能够通过计算预测和实验确认对（-）可卡因具有进一步提高的催化活性的新型人 BChE 突变体，包括最活跃的突变体（A199S/F227S/S287G/A328W/Y332G 突变体），与野生型人 BChE 相比，催化效率提高了 2790 倍（/ = 2.5 × 10 min M）。与在报道的可卡因依赖治疗酶疗法的临床开发中测试的参考突变体（A199S/S287G/A328W/Y332G 突变体）相比，这种新型突变体（预测有新的 F227S 突变）对（-）可卡因的催化效率提高了约 2.6 倍。计算和实验 ln（RCE）值之间的良好一致性表明，获得的相关性对于计算预测是稳健的。在研究具有氧阴离子穴的 BChE 或其他丝氨酸水解酶/酯酶时，也可以探索类似的相关性，以稳定羰基氧在酶水解其他底物的速率决定步骤中的氧阴离子穴。
