INTEGRATION OF F1 AND F0 ECTORS OF E. COLI PROTON-ATPASE

“b”亚基(也称为“uncf”蛋白质)是蛋白质的三个亚基之一。 大肠杆菌质子-ATPase的FO区。其氨基酸序列已知。 最近的数据表明亚单位b与F1-扇区( ATP合成/水解催化单元)到FO-部门( 膜中的质子通道)。综合来看，对 B亚基可能的二级和三级结构，证据 表明这个亚基在F1的整合中起着核心作用 和Fo扇区以及在ATP水解与H+离子转运的耦合中 (“能量耦合”)。 为了评估特定残基和结构域的重要性 亚基b在F1-Fo积分中，亚基b将被各种 包括化学修饰、有限蛋白水解法、本地化技术 诱变。将通过分析影响来建立功能关联 B亚基修饰对被动质子通过雾的传输的影响 特异的F1结合、ATP驱动的质子泵浦和传递 质子梯度诱导的F1构象变化。交联性 将进行实验以确定亚基的哪些残基或结构域 B与F1-亚基结合，部分逆转株的检测将 作为发现亚基间相互作用的另一种方式进行 在b亚基和质子-ATPase的其他亚基之间。 离子转运ATPase的“能量耦合”本质是一种 生物学的基本问题。通过集中于亚单位b，我们将推导出 对这一过程的分子洞察。

Subunit "b" (also called "uncF" protein) is one of three subunits in the Fo-sector of E. coli proton-ATPase. Its amino acid sequence is known. Recent data implicates subunit b in specific binding of the F1-sector (the ATP synthesis/hydrolysis catalytic unit) to the Fo-sector (the proton-channel in the membrane). Taken together with predictions of the likely secondary and tertiary structure of subunit b, the evidence indicates that this subunit plays a central role in integration of the F1 and Fo sectors and in the coupling of ATP hydrolysis to H+ ion transport ("energy coupling"). In order to assess the importance of specific residues and domains of subunit b in F1-Fo integration, subunit b will be modified by a variety of techniques including chemical modification, limited proteolysis, localized mutagenesis. Functional correlations will be made by assaying the effects of the subunit b modifications on passive proton-transport through Fo specific F1-binding, ATP-driven proton-pumping and transmittal of proton-gradient-induced conformational changes to F1. Cross-linking experiments will be done to determine which residues or domains of subunit b bind to F1-subunits, and examination of partial revertants will be carried out as an alternative way of uncovering intersubunit interations between subunit b and other subunits of the proton-ATPase. The nature of "energy-coupling" in ion-transporting ATPases is a fundamental problem of biology. By focusing on subunit b we will deduce molecular insights into this process.