Mandel M, Moriyama Y, Hulmes J D, Pan Y C, Nelson H, Nelson N
Roche Institute of Molecular Biology, Roche Research Center, Nutley, NJ 07110.
Proc Natl Acad Sci U S A. 1988 Aug;85(15):5521-4. doi: 10.1073/pnas.85.15.5521.
Vacuolar H+-ATPases function in generating protonmotive force across the membranes of organelles connected with the vacuolar system of eukaryotic cells. This family of H+-ATPases is distinct from the two other families of H+-ATPases, the plasma membrane-type and the eubacterial-type. One of the subunits of the vacuolar H+-ATPase binds N,N'-dicyclohexylcarbodiimide (DCCD) and has been implicated in the proton-conducting activity of these enzymes. We have cloned and sequenced the gene encoding the DCCD-binding protein (proteolipid) of the H+-ATPase of bovine chromaffin granules. The gene encodes a highly hydrophobic protein of 15,849 Da. Hydropathy plots revealed four transmembrane segments, one of which contains a glutamic residue that is the likely candidate for the DCCD binding site. Sequence homology with the vacuolar proteolipid and with the proteolipids of eubacterial-type H+-ATPases was detected. The proteolipids from Escherichia coli, spinach chloroplasts, and yeast mitochondria matched better to the NH2-terminal part of the vacuolar protein. The proteolipids of bovine mitochondria and Neurospora mitochondria matched better to the COOH-terminal end of the vacuolar proteolipid. These findings suggest that the proteolipids of the vacuolar H+-ATPases were evolved in parallel with the eubacterial proteolipid, from a common ancestral gene that underwent gene duplication.
液泡H⁺-ATP酶的功能是在与真核细胞液泡系统相连的细胞器膜上产生质子动力。该H⁺-ATP酶家族不同于另外两个H⁺-ATP酶家族，即质膜型和真细菌型。液泡H⁺-ATP酶的一个亚基能结合N,N'-二环己基碳二亚胺（DCCD），并与这些酶的质子传导活性有关。我们已经克隆并测序了牛嗜铬粒蛋白H⁺-ATP酶的DCCD结合蛋白（蛋白脂质）的编码基因。该基因编码一个15849Da的高度疏水蛋白。亲水性图谱显示有四个跨膜片段，其中一个含有谷氨酸残基，这可能是DCCD结合位点的候选者。检测到与液泡蛋白脂质和真细菌型H⁺-ATP酶的蛋白脂质的序列同源性。来自大肠杆菌、菠菜叶绿体和酵母线粒体的蛋白脂质与液泡蛋白的氨基末端部分匹配得更好。牛线粒体和粗糙脉孢菌线粒体的蛋白脂质与液泡蛋白脂质的羧基末端匹配得更好。这些发现表明，液泡H⁺-ATP酶的蛋白脂质与真细菌蛋白脂质是从一个经历了基因复制的共同祖先基因平行进化而来的。
