de Mendonça R L, Beck E, Rumjanek F D, Goffeau A
Université Catholique de Louvain, Unité de Biochimie Physiologique, Louvain-la-Neuve, Belgium.
Mol Biochem Parasitol. 1995 Jun;72(1-2):129-39. doi: 10.1016/0166-6851(95)00078-f.
Complementary DNA was isolated, encoding a putative Ca(2+)-transport ATPase (SMA1) of the human parasitic trematode Schistosoma mansoni. The cDNA was isolated by a nested polymerase chain reaction based strategy. The oligonucleotides used were designed on the basis of conserved amino-acid regions found in P-type ATPases. The complete nucleotide sequence was determined. The primary structure and topology of the enzyme were deduced. SMA1 has 1022 amino acids and a predicted molecular mass of 113 kDa. This protein is 67% identical and phylogenetically related to several sarco/endoplasmic reticulum Ca(2+)-ATPases but lacks the phospholamban-binding domain that exists in the SERCA isoforms 1 and 2. The membrane topology predicted for SMA1 is characteristic of the P-type ATPases, showing two major cytoplasmic loops and ten conserved hydrophobic segments. Sequences and residues that are important for the function of the SER Ca(2+)-ATPase, such as the high-affinity Ca(2+)-binding sites, the putative fluorescein isothiocyanate binding site, the 5'-(p-fluorosulfonyl)benzoyladenosine binding site and the aspartyl phosphorylation site, are conserved in SMA1, suggesting that the cloned gene is a Ca(2+)-transport ATPase of the SERCA family. In addition, three PCR products were cloned which share homology with another SER Ca(2+)-ATPase, with the yeast secretory pathway Ca(2+)-ATPase PMR1 and its mammalian homologue, and with the alpha subunit of a Na+,K(+)-ATPase.
分离出了互补DNA，其编码曼氏血吸虫这种人体寄生吸虫的一种假定的Ca(2+)转运ATP酶（SMA1）。该cDNA是通过基于巢式聚合酶链反应的策略分离得到的。所用的寡核苷酸是根据在P型ATP酶中发现的保守氨基酸区域设计的。确定了完整的核苷酸序列。推导了该酶的一级结构和拓扑结构。SMA1有1022个氨基酸，预测分子量为113 kDa。该蛋白质与几种肌质/内质网Ca(2+) - ATP酶有67%的同一性且在系统发育上相关，但缺乏存在于肌质/内质网Ca(2+) - ATP酶同工型1和2中的受磷蛋白结合结构域。预测的SMA1膜拓扑结构是P型ATP酶的特征，显示出两个主要的胞质环和十个保守的疏水片段。对于肌质网Ca(2+) - ATP酶功能重要的序列和残基（如高亲和力Ca(2+)结合位点、假定的异硫氰酸荧光素结合位点、5'-(对氟磺酰基)苯甲酰腺苷结合位点和天冬氨酰磷酸化位点）在SMA1中是保守的，这表明克隆的基因是肌质/内质网Ca(2+) - ATP酶家族中的一种Ca(2+)转运ATP酶。此外，克隆了三个PCR产物，它们与另一种肌质网Ca(2+) - ATP酶、与酵母分泌途径Ca(2+) - ATP酶PMR1及其哺乳动物同源物以及与Na+,K(+) - ATP酶的α亚基具有同源性。
