Inoue Takao, Wang Yanru, Jefferies Kevin, Qi Jie, Hinton Ayana, Forgac Michael
Department of Physiology, Tufts University School of Medicine, 136 Harrison Ave., Boston, Massachusetts 02111, USA.
J Bioenerg Biomembr. 2005 Dec;37(6):393-8. doi: 10.1007/s10863-005-9478-8.
The V-ATPases are ATP-dependent proton pumps present in both intracellular compartments and the plasma membrane. They function in such processes as membrane traffic, protein degradation, renal acidification, bone resorption and tumor metastasis. The V-ATPases are composed of a peripheral V(1) domain responsible for ATP hydrolysis and an integral V(0) domain that carries out proton transport. Our recent work has focused on structural analysis of the V-ATPase complex using both cysteine-mediated cross-linking and electron microscopy. For cross-linking studies, unique cysteine residues were introduced into structurally defined sites within the B and C subunits and used as points of attachment for the photoactivated cross-linking reagent MBP. Disulfide mediated cross-linking has also been used to define helical contact surfaces between subunits within the integral V(0) domain. With respect to regulation of V-ATPase activity, we have investigated the role that intracellular environment, luminal pH and a unique domain of the catalytic A subunit play in controlling reversible dissociation in vivo.
V-ATP酶是一种依赖ATP的质子泵，存在于细胞内区室和质膜中。它们在膜运输、蛋白质降解、肾脏酸化、骨吸收和肿瘤转移等过程中发挥作用。V-ATP酶由负责ATP水解的外周V(1)结构域和进行质子运输的整合V(0)结构域组成。我们最近的工作集中在使用半胱氨酸介导的交联和电子显微镜对V-ATP酶复合物进行结构分析。对于交联研究，将独特的半胱氨酸残基引入B和C亚基内结构明确的位点，并用作光活化交联试剂MBP的附着点。二硫键介导的交联也已用于确定整合V(0)结构域内亚基之间的螺旋接触表面。关于V-ATP酶活性的调节，我们研究了细胞内环境、管腔pH值和催化A亚基的一个独特结构域在体内控制可逆解离中所起的作用。
