Centre of Membrane Proteins and Receptors (COMPARE), University of Nottingham, Midlands, UK.
Division of Physiology, Pharmacology & Neuroscience, School of Life Sciences, University of Nottingham, Nottingham, UK.
Pharmacol Res Perspect. 2022 Aug;10(4):e00978. doi: 10.1002/prp2.978.
In this study, we report the β -adrenoceptor binding kinetics of several clinically relevant β -adrenoceptor (β AR) agonists and antagonists. [ H]-DHA was used to label CHO-β AR for binding studies. The kinetics of ligand binding was assessed using a competition association binding method. Ligand physicochemical properties, including logD and the immobilized artificial membrane partition coefficient (K ), were assessed using column-based methods. Protein Data Bank (PDB) structures and hydrophobic and electrostatic surface maps were constructed in PyMOL. We demonstrate that the hydrophobic properties of a molecule directly affect its kinetic association rate (k ) and affinity for the β AR. In contrast to our findings at the β -adrenoceptor, K , reflecting both hydrophobic and electrostatic interactions of the drug with the charged surface of biological membranes, was no better predictor than simple hydrophobicity measurements such as clogP or logD , at predicting association rate. Bisoprolol proved kinetically selective for the β AR subtype, dissociating 50 times slower and partly explaining its higher measured affinity for the β AR. We speculate that the association of positively charged ligands at the β AR is curtailed somewhat by its predominantly neutral/positive charged extracellular surface. Consequently, hydrophobic interactions in the ligand-binding pocket dominate the kinetics of ligand binding. In comparison at the β AR, a combination of hydrophobicity and negative charge attracts basic, positively charged ligands to the receptor's surface promoting the kinetics of ligand binding. Additionally, we reveal the potential role kinetics plays in the on-target and off-target pharmacology of clinically used β-blockers.
在这项研究中，我们报告了几种临床相关β-肾上腺素受体（βAR）激动剂和拮抗剂的β-肾上腺素受体结合动力学。[H]-DHA 用于标记 CHO-βAR 进行结合研究。使用竞争关联结合方法评估配体结合的动力学。使用基于柱的方法评估配体的物理化学性质，包括 logD 和固定化人工膜分配系数（K）。使用 PyMOL 构建蛋白质数据库（PDB）结构和疏水性和静电表面图。我们证明分子的疏水性直接影响其动力学缔合速率（k）和与βAR 的亲和力。与我们在β-肾上腺素受体上的发现相反，K 反映了药物与生物膜带电表面的疏水性和静电相互作用，与简单的疏水性测量（如 clogP 或 logD）相比，不能更好地预测缔合速率。比索洛尔在动力学上对βAR 亚型具有选择性，解离速度慢 50 倍，部分解释了其对βAR 的更高测量亲和力。我们推测，带正电荷的配体在βAR 上的结合由于其主要带中性/正电荷的细胞外表面而受到一定程度的限制。因此，配体结合口袋中的疏水相互作用主导了配体结合的动力学。相比之下，βAR 上的疏水性和负电荷结合吸引碱性、带正电荷的配体到受体表面，促进配体结合的动力学。此外，我们揭示了动力学在临床上使用的β阻滞剂的靶内和靶外药理学中的潜在作用。
