Structure of human anti-cancer drug resistance by hABCG2

描述(申请人提供)：人类ATP结合盒乳腺癌耐药蛋白BCRP(ABCG2)的自发突变通过输出经典的抗癌药物来传递肿瘤细胞耐药性，通常会导致致命的化疗复发。人ABCG2是一种同源二聚体的完整膜蛋白，具有独特而紧凑的“半转运体”设计，不需要任何糖基化就可以起到药物出口泵的作用。这些特性可能使该蛋白成为用于高分辨结构测定的人类多药输出泵的最佳候选者。从巴斯德毕赤酵母中表达低毫克量ABCG2的方案已经开发出来。既然已经确定了稳定hABCG2的非离子洗涤剂，那么从无细胞系统中生产更高数量的蛋白质将得到优化。这项提议的目标是通过X射线结晶学解决人类ABCG2的结构，以可视化这种临床上重要的蛋白质将抗癌药物赶出细胞的机制。“点击”化学技术将被应用于纯化的ABCG2蛋白质和蛋白质晶体，以识别这种药物出口泵的特定抑制剂的构建块。一种高分辨率的晶体结构将与“点击”化学的结果相结合，以实现下一代抗癌药物的设计，这种药物将专门阻断ABCG2的活性，并提高对几种人类癌症的化疗效果。

DESCRIPTION (provided by applicant): Spontaneous mutations of the human ATP-binding cassette breast cancer resistance protein, BCRP (ABCG2), convey tumor cell resistance by exporting classical anti-cancer agents, often resulting in a fatal relapse of chemotherapy. Human ABCG2 is a homodimeric integral membrane protein with a unique and compact "half-transporter" design and does not require any glycosylation to function as a drug export pump. These characteristics may make this protein the best candidate among human multidrug export pumps for high-resolution structure determination. The protocols for the expression of low milligram quantities of ABCG2 from the yeast Pichia pastoris have already been developed. Production of even higher quantities of protein from cell-free systems will be optimized now that non-ionic detergents that stabilize hABCG2 have already been identified. The goal of this proposal is to solve the structure of human ABCG2 by x-ray crystallography to visualize the mechanisms by which this clinically important protein drives anti-cancer pharmaceuticals out of cells. "Click" chemistry techniques will be applied to purified ABCG2 protein and protein crystals to identify building blocks of specific inhibitors to this drug export pump. A high-resolution crystal structure will be used in combination with the output from the "click" chemistry to enable the design of next-generation anti-cancer pharmaceuticals that will specifically block ABCG2 activity and increase the effectiveness of chemotherapy in several human cancers.