A novel signal transduction mechanism via intracellular ATP sensor

atp结合盒（ABC）蛋白包括一个结构相关的膜蛋白家族，共享保守的核苷酸结合域。它们包括MDR1，一种atp依赖性外排泵，可挤出有毒的异种生物，囊性纤维化跨膜传导调节剂（CFTR），阴离子通道，磺酰脲受体（SUR 1），胰腺atp敏感的K^+通道调节剂。本研究的目的是：(1)深入了解ABC蛋白的转运体、通道和调节因子的分子机制。2)了解具有双重功能的分子机制。3)探讨ABC蛋白的生理功能在细胞信号转导中的作用机制。胰岛β细胞中的K_<ATP>通道在调节葡萄糖诱导的胰岛素分泌中起关键作用。虽然电生理研究为ATP、MgADP和药物对K_<ATP>通道的复杂调控提供了线索，但更多的K_<ATP>通道调控的分子机制尚不清楚。K_<ATP>通道是ABC超家族SUR 1亚基的异寡聚复合物，具有两个核苷酸结合折叠和形成孔的Kir6.2亚基。我们发现MgATP和MgADP稳定了8-叠氮-[α -^<32>P]ATP与SUIR1的结合，而不是MgATP-gamma。SUIR1的NBF2的Walker A和B基序突变消除了MgADP的这种稳定作用。这些结果表明，SURl在NBF1上强烈结合8-叠氮- atp，而MgADP通过直接结合NBF2或在NBF2上水解结合的MgATP，稳定了NBF1上预结合的8-叠氮- atp结合。磺脲类格列苯脲在MgADP或MgATP存在的情况下，以浓度依赖性的方式从SUR1释放预结合的8-叠氮-[α -^<32>P]ATP。这一直接的生化证据表明，在SUR1的两个NBFs的核苷酸结合中，格列本脲都阻断了ATP和MgADP的合作结合，并与MgADP结合在NBF2上，导致ATP从NBF1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