Department of Medicinal Chemistry, University of Washington, Seattle, Washington 98195-7610, United States.
Biochemistry. 2022 Apr 19;61(8):730-740. doi: 10.1021/acs.biochem.2c00056. Epub 2022 Apr 6.
The ABC efflux pump P-glycoprotein (P-gp) transports a wide variety of drugs and is inhibited by others. Some drugs stimulate ATP hydrolysis at the nucleotide binding domains (NBDs) and are transported, others uncouple ATP hydrolysis and transport, and others inhibit ATP hydrolysis. The molecular basis for the different behavior of these drugs is not well understood despite the availability of several structural models of P-gp complexes with ligands bound. Hypothetically, ligands differentially alter the conformational dynamics of peptide segments that mediate the coupling between the drug binding sites and the NBDs. Here, we explore by hydrogen-deuterium exchange mass spectrometry the dynamic consequences of a classic substrate and inhibitor, vinblastine and zosuquidar, binding to mouse P-gp (mdr1a) in lipid nanodiscs. The dynamics of P-gp in nucleotide-free, pre-hydrolysis, and post-hydrolysis states in the presence of each drug reveal distinct mechanisms of ATPase stimulation and implications for transport. For both drugs, there are common regions affected in a similar manner, suggesting that particular networks are the key to stimulating ATP hydrolysis. However, drug binding effects diverge in the post-hydrolysis state, particularly in the intracellular helices (ICHs 3 and 4) and neighboring transmembrane helices. The local dynamics and conformational equilibria in this region are critical for the coupling of drug binding and ATP hydrolysis and are differentially modulated in the catalytic cycle.
ABC 外排泵 P-糖蛋白 (P-gp) 可转运多种药物，并受到其他药物的抑制。一些药物可刺激核苷酸结合域（NBD）的 ATP 水解并进行转运，另一些药物则使 ATP 水解与转运解偶联，还有一些药物则抑制 ATP 水解。尽管已有几种与配体结合的 P-gp 复合物的结构模型，但这些药物的不同行为的分子基础仍未得到很好的理解。理论上，配体可改变介导药物结合位点与 NBD 之间偶联的肽段的构象动力学。在这里，我们通过氘氢交换质谱法研究了经典底物长春碱和抑制剂唑舒达与脂质纳米盘中的鼠 P-gp（mdr1a）结合时的动态后果。在每种药物存在的情况下，无核苷酸、预水解和水解后状态下的 P-gp 的动力学揭示了 ATP 酶刺激的不同机制，并对转运具有影响。对于这两种药物，有共同的受影响区域以相似的方式受到影响，这表明特定的网络是刺激 ATP 水解的关键。然而，在水解后状态下，药物结合的影响存在差异，特别是在细胞内螺旋（ICHs 3 和 4）和相邻的跨膜螺旋中。该区域的局部动力学和构象平衡对于药物结合和 ATP 水解的偶联至关重要，并且在催化循环中受到不同的调节。
