Ion transport by the human Breast Cancer Resistance Protein (ABCG2)

药物转运体，如乳腺癌耐药蛋白（ABCG2），被认为是人类药物分布的关键参与者。这种外排泵在负责药物生物转化和排泄的器官中的定位使ABCG2在控制药物进入代谢酶和排泄途径方面具有重要的守门人功能。在癌细胞中也发现了ABCG2，它介导药物向细胞外部的挤压。这样一来，它就阻止了抗癌药物进入细胞，从而削弱了对这种危及生命的疾病的化疗治疗。在本提案中，我们将研究人类ABCG2转运机制的基本方面。特别是，我们将研究以前未知的ABCG2传输离子（如质子）的能力。我们有兴趣了解哪些离子被运输（除了质子，例如钠、钾、氯离子），ABCG2是如何运输这些离子的，为什么？离子输运和药物输运之间有什么关系？ABCG2被认为作为二聚体具有活性，我们将通过质谱法进行测试，并且可能具有药物结合位点，交替暴露于膜的内表面（药物结合的地方）和膜的外表面（药物释放的地方）。基于多药物结合蛋白（转运蛋白和转录调节蛋白）的晶体结构，我们假设(i)质子和其他离子可能在药物释放过程中将药物从ABCG2的结合位点上取代，或者（ii）它们的结合可能支持两个ABCG2单元及其界面的结构变化，这些变化与药物结合位点的重新定向有关。关于ABCG2活性的基础知识将允许合理开发这种外排泵的抑制剂（也称为调节剂），可用于将药物靶向人体特定部位，并改善癌症的化疗。

Drug transporters, such as the Breast Cancer Resistance Protein (ABCG2), are recognized as key players in the distribution of drugs in human. The localization of this efflux pump in organs responsible for drug biotransformation and excretion gives ABCG2 an important gatekeeper function in controlling drug access to metabolizing enzymes and excretory pathways. ABCG2 is also found in cancer cells, where it mediates the extrusion of drug to the cell's exterior. In doing so, it prevents entry of anticancer drugs into cells and, hence, impairs the chemotherapeutic treatment of this life-threatening disease. In this proposal, we will study fundamental aspects of the transport mechanism of human ABCG2. In particular, we will study the previously unknown ability of ABCG2 to transport ions such as protons. We are interested to learn which ions are transported (in addition to protons, for example sodium, potassium, chloride ions), how ABCG2 transports these ions, and why? What is the relationship between ion transport and drug transport? ABCG2 is thought to be active as a dimer, which we will test by mass spectrometry, and is likely to have drug binding sites that are alternately exposed to the inside surface of the membrane (where drugs bind) and outside surface of the membrane (where drugs are released). Based on the available crystal structures of multidrug binding proteins (transporters and transcriptional regulators), we hypothesize that (i) protons and other ions might displace drugs from binding sites in ABCG2 during drug release and/or (ii) their binding might support structural changes in the two ABCG2 units and their interface, that are associated with the reorientation of the drug binding sites. Fundamental knowledge about ABCG2 activity will allow the rational development of inhibitors (also termed modulators) of this efflux pump that could be used to target drugs to specific parts of the human body, and to improve chemotherapy of cancers.