Drug-Induced Conformational Dynamics of P-Glycoprotein Underlies the Transport of Camptothecin Analogs.
Drug-Induced Conformational Dynamics of P-Glycoprotein Underlies the Transport of Camptothecin Analogs.
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DOI：
10.3390/ijms242216058
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发表时间：
2023-11-07
影响因子：
5.6
通讯作者：
中科院分区：
文献类型：
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P-糖蛋白(Pgp)在药物的生物利用度和多药耐药的形成中起着关键作用。了解蛋白质的活性和设计有效的药物需要深入了解PGP介导的外源物质运输的潜在机制。在这项研究中，我们研究了药物诱导的Pgp构象变化，并采用构象门控模型来解释Pgp介导的喜树碱类似物(CPT)的转运。虽然PGP显示了广泛的构象，但我们将其简化为三种模型状态：“向内开放”、“向外开放”和“中间”。利用丙烯酰胺猝灭PGP荧光作为检测蛋白质三级结构的工具，我们观察到拓扑替康(TPT)、SN-38和伊立替康(IRT)诱导蛋白质发生明显的构象变化。TPT引起了类似于AMPPNP的重大转变，暗示着ATP不依赖于“开放-向外”构象。IRT和SN-38对Pgp构象的影响相对较小。实验原子力显微镜(AFM)成像支持这些发现。此外，ATPase的水解率与配体诱导的PGP构象变化有关。我们假设核苷酸结合域(NBD)之间的分离为底物运输创建了构象屏障。减少构象障碍的底物，如TPT，可以更好地运输。从PGP介导的ATP水解动力学曲线中提取的TPT对ATP的亲和力分别比SN-38和IRT高约2倍和3倍。反之，荧光猝灭法测定的解离常数(Kd)无显著差异。TPT和IRT与PGP的饱和转移双差(STDD)核磁共振谱表明，PGP与CPTS相互作用的原因是相似的官能团。在现有的CPT和DNA-拓扑异构酶-I复合体结构-活性关系数据的指导下，旨在修饰这些官能团的努力可能为开发更有效的下一代CPT铺平道路。
P-glycoprotein (Pgp) plays a pivotal role in drug bioavailability and multi-drug resistance development. Understanding the protein’s activity and designing effective drugs require insight into the mechanisms underlying Pgp-mediated transport of xenobiotics. In this study, we investigated the drug-induced conformational changes in Pgp and adopted a conformationally-gated model to elucidate the Pgp-mediated transport of camptothecin analogs (CPTs). While Pgp displays a wide range of conformations, we simplified it into three model states: ‘open-inward’, ‘open-outward’, and ‘intermediate’. Utilizing acrylamide quenching of Pgp fluorescence as a tool to examine the protein’s tertiary structure, we observed that topotecan (TPT), SN-38, and irinotecan (IRT) induced distinct conformational shifts in the protein. TPT caused a substantial shift akin to AMPPNP, suggesting ATP-independent ‘open-outward’ conformation. IRT and SN-38 had relatively moderate effects on the conformation of Pgp. Experimental atomic force microscopy (AFM) imaging supports these findings. Further, the rate of ATPase hydrolysis was correlated with ligand-induced Pgp conformational changes. We hypothesize that the separation between the nucleotide-binding domains (NBDs) creates a conformational barrier for substrate transport. Substrates that reduce the conformational barrier, like TPT, are better transported. The affinity for ATP extracted from Pgp-mediated ATP hydrolysis kinetics curves for TPT was about 2-fold and 3-fold higher than SN-38 and IRT, respectively. On the contrary, the dissociation constants (KD) determined by fluorescence quenching for these drugs were not significantly different. Saturation transfer double difference (STDD) NMR of TPT and IRT with Pgp revealed that similar functional groups of the CPTs are accountable for Pgp-CPTs interactions. Efforts aimed at modifying these functional groups, guided by available structure-activity relationship data for CPTs and DNA-Topoisomerase-I complexes, could pave the way for the development of more potent next-generation CPTs.
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影响因子：
19.6
作者：
Anglicheau, D.;Pallet, N.;Thervet, E.
通讯作者：
Thervet, E.
DOI：
10.1016/j.jphotochem.2012.08.015
发表时间：
2012-12-01
影响因子：
4.3
作者：
Campbell, Katie;Zappas, Andrew;Bucknall, David G.
通讯作者：
Bucknall, David G.
影响因子：
4.8
作者：
Eckford, Paul D. W.;Sharom, Frances J.
通讯作者：
Sharom, Frances J.
影响因子：
--
作者：
Ando K;Shah AK;Sachdev V;Kleinstiver BP;Taylor-Parker J;Welch MM;Hu Y;Salgia R;White FM;Parvin JD;Ozonoff A;Rameh LE;Joung JK;Bharti AK
通讯作者：
Bharti AK
影响因子：
3.7
作者：
Patra M;Mukhopadhyay C;Chakrabarti A
通讯作者：
Chakrabarti A