Study on ion-coupled subunit interaction of Na-HransbcatingV-ATPase

(I)河肠球菌液泡型(V型)钠离子泵送三磷酸腺苷酶(Na+-ATPase)的膜转子环由10个NtpK亚基组成，它们分别是其他V-ATPase和FIFO-或F-ATPase中的16千和8千道顿蛋白脂的同源物。每个NtpK亚基都有四个跨膜α螺旋，在螺旋2和螺旋4之间有一个钠离子结合，这个位置深埋在膜中，包括基本残基谷氨酸-139。这个位置可能通过NtpI亚基中的两个半通道连接到膜表面，环相对于NtpI亚基旋转。转化区和催化区的对称性不匹配似乎是V-ATPase和F-ATPase的内在特征。(Ii)通过对NTP基因缺失突变株形成V-ATPase复合体的生化分析，我们发现VI部分由亚基I和K的A、B、C、D、E、F、G和V0部分组成，B亚基不是形成ACDIK复合体所必需的，A亚基在V0V1复合体的形成中起关键作用。这些结果对于了解V-ATPase亚基在离子转运过程中的分子相互作用具有重要意义。

(i) The membrane rotor ring from the vacuolar-type (V-type) sodium ion-pumping adenosine triphosphatase (Na+-ATPase) from Enterococcus hirae consists of 10 NtpK subunits, which are homologs of the 16-kilodalton and 8-kilodalton proteolipids found in other V-ATPases and in FIFo-or F-ATPases, respectively. Each NtpK subunit has four transmembrane alpha helices, with a sodium ion bound between helices 2 and 4 at a site buried deeply in the membrane that includes the essential residue glutamate-139. This site is probably connected to the membrane surface by two half-channels in subunit NtpI, against which the ring rotates. Symmetry mismatch between the rotor and catalytic domains appears to be an intrinsic feature of both V-and F-ATPases.(ii) By biochemical analysis of V-ATPase complex formation with the ntp gene-deleted mutant strains, we found that the VI portion is composed of subunit A, B, C, D, E, F, G and V0 portion of subunit I and K. Subunit B is not necessary to form the ACDIK complex, and subunit A plays a key role for V0V1 complex formation. All these results are important for understanding the molecular interaction of V-ATPase subunits during ion translocation.