A novel signal transduction mechanism via intracellular ATP sensor

ATP结合盒（ABC）蛋白是一类结构上相关的膜蛋白家族，具有高度保守的核苷酸结合域。包括MDR 1（一种ATP依赖的外排泵，可排出有毒的外源性物质）、囊性纤维化跨膜传导调节因子（CFTR）（一种阴离子通道）和磺酰脲受体（SUR 1）（胰腺ATP敏感的K^+通道调节因子）。研究ABC蛋白的生理功能及其调节因子; 2）了解ABC蛋白具有双重功能的分子机制; 3）探讨ABC蛋白在细胞信号转导机制中的生理功能<ATP>。虽然电生理学研究为ATP、MgADP和药物对钾通道的复杂调控提供了线索<ATP>，但其分子机制尚不清楚。 ...更多信息 钾通道的<ATP>调节机制尚不清楚。K_<ATP>通道是由ABC超家族的SUR 1亚基与成孔性Kir6.2亚基形成的异源寡聚复合物，具有两个核苷酸结合折叠。我们发现MgATP和MgADP，而不是MgATP-γ S，稳定了预结合的8-叠氮基-[α-^<32>P]ATP与SUIR 1的结合。SUIR 1的NBF 2的步行者A和B基序的突变消除了MgADP的这种稳定作用。这些结果表明，SUR 1在NBF 1处强烈地结合8-叠氮基-ATP，并且MgADP通过直接结合NBF 2或水解结合在NBF 2处的MgATP来稳定NBF 1处的预先结合的8-叠氮基-ATP结合。磺酰脲类药物格列本脲在MgADP或MgATP存在下以浓度依赖性方式引起SUR 1释放预结合的8-叠氮基-[α-^<32>P]ATP。SUR 1的两个NBF核苷酸结合中协同相互作用的直接生化证据表明，格列本脲既阻断ATP和MgADP的这种协同结合，又与NBF 2结合的MgADP协同作用，导致ATP从NBF 1释放。少

ATP-binding cassette (ABC) proteins comprise a family of structurally related membrane proteins sharing well conserved nucleotide binding domains. They include MDR1, an ATP-dependent efflux pump that extrude toxic xenobiotics, the cystic fibrosis transmembrane conductance regulator (CFTR), an anion channel, and the sulfonylurea receptor (SUR 1), the pancreatic ATP-sensitive K^+ channel regulator.The objective of the research was1) to develop a deep understanding of the molecular mechanisms of transporters, channels, and regulators of ABC proteins.2) to understand the molecular mechanisms of having dual functions.3) to explore the physiological functions of ABC proteins in cellular signal transduction mechanism.The K_<ATP> channels in pancreatic beta-cells are critical in the regulation of glucose-induced insulin secretion. Although electrophysiological studies provide clues to the complex control of K_<ATP> channels by ATP, MgADP, and the pharmacological agents, the molecular mechanism … More of K_<ATP> channel regulation remains unclear. The K_<ATP> channel is a hetero-oligomeric complex of SUR 1 subunits of ABC superfamily with two nucleotide- binding folds and the pore-forming Kir6.2 subunits. We revealed that MgATP and MgADP, but not MgATP-gammaS, stabilize binding of prebound 8-azido-[alpha-^<32>P]ATP to SUIR1. Mutation in the Walker A and B motifs of NBF2 of SUIR1 abolished this stabilizing effect of MgADP.These results suggest that SURl binds 8-azido-ATP strongly at NBF1, and that MgADP, either by direct binding to NBF2 or hydrolysis of bound MgATP at NBF2, stabilizes prebound 8-azido-ATP binding at NBF1. The sulfonylurea glibenclamide caused release of prebound 8-azido-[alpha-^<32>P]ATP from SUR1 in the presence of MgADP or MgATP in a concentration-dependent manner. This direct biochemical evidence of cooperative interaction in nucleotide binding of the two NBFs of SUR1 suggests that glibenclamide both blocks this cooperative binding of ATP and MgADP, and, in cooperation with the MgADP bound at NBF2, causes ATP to be released from NBF1. Less