Hou W-R, Hou Y-L, Ding X, Wang T
Key Laboratory of Southwest China Wildlife Resources Conservation, Ministry of Education, College of Life Science, China West Normal University, P.R. China.
Genet Mol Res. 2012 Sep 3;11(3):3164-74. doi: 10.4238/2012.September.3.5.
The ATP5G1 gene is one of the three genes that encode mitochondrial ATP synthase subunit c of the proton channel. We cloned the cDNA and determined the genomic sequence of the ATP5G1 gene from the giant panda (Ailuropoda melanoleuca) using RT-PCR technology and touchdown-PCR, respectively. The cloned cDNA fragment contains an open reading frame of 411 bp encoding 136 amino acids; the length of the genomic sequence is of 1838 bp, containing three exons and two introns. Alignment analysis revealed that the nucleotide sequence and the deduced protein sequence are highly conserved compared to Homo sapiens, Mus musculus, Rattus norvegicus, Bos taurus, and Sus scrofa. The homologies for nucleotide sequences of the giant panda ATP5G1 to those of these species are 93.92, 92.21, 92.46, 93.67, and 92.46%, respectively, and the homologies for amino acid sequences are 90.44, 95.59, 93.38, 94.12, and 91.91%, respectively. Topology prediction showed that there is one protein kinase C phosphorylation site, one casein kinase II phosphorylation site, five N-myristoylation sites, and one ATP synthase c subunit signature in the ATP5G1 protein of the giant panda. The cDNA of ATP5G1 was transfected into Escherichia coli, and the ATP5G1 fused with the N-terminally GST-tagged protein gave rise to accumulation of an expected 40-kDa polypeptide, which had the characteristics of the predicted protein.
ATP5G1基因是编码质子通道线粒体ATP合酶亚基c的三个基因之一。我们分别使用RT-PCR技术和降落PCR从大熊猫（Ailuropoda melanoleuca）中克隆了cDNA并确定了ATP5G1基因的基因组序列。克隆的cDNA片段包含一个411 bp的开放阅读框，编码136个氨基酸；基因组序列长度为1838 bp，包含三个外显子和两个内含子。比对分析表明，与智人、小家鼠、褐家鼠、牛和野猪相比，该核苷酸序列和推导的蛋白质序列高度保守。大熊猫ATP5G1基因核苷酸序列与这些物种的同源性分别为93.92%、92.21%、92.46%、93.67%和92.46%，氨基酸序列的同源性分别为90.44%、95.59%、93.38%、94.12%和91.91%。拓扑预测显示，大熊猫ATP5G1蛋白中有一个蛋白激酶C磷酸化位点、一个酪蛋白激酶II磷酸化位点、五个N-肉豆蔻酰化位点和一个ATP合酶c亚基特征序列。将ATP5G1的cDNA转染到大肠杆菌中，与N端带有GST标签的蛋白融合的ATP5G1导致预期的40 kDa多肽积累，该多肽具有预测蛋白的特征。
