Rickers J, Cushman J C, Michalowski C B, Schmitt J M, Bohnert H J
Department of Biochemistry, University of Arizona, Tucson 85721.
Mol Gen Genet. 1989 Feb;215(3):447-54. doi: 10.1007/BF00427042.
We have determined the complete nucleotide sequence of a full length cDNA encoding the Crassulacean acid metabolism (CAM) isogene of phospho(enol)pyruvate carboxylase (PEPCase). The cDNA clone, 3348 bp in length, was obtained from mRNA isolated from Mesembryanthemum crystallinum (common ice plant) which had undergone salt stress and subsequent induction of CAM. The long open reading frame encodes PEPCase (EC 4.1.1.31) with a predicted molecular mass of 110533 daltons. The deduced amino acid sequence of the ice plant PEPCase is most similar to that from maize having an amino acid identity of 74.9%. Sequence identity in corresponding regions of the PEPCase proteins from Escherichia coli and the cyanobacterium Anacystis nidulans are 41.4% and 33.5%, respectively. A compilation of the four amino acid sequences permitted the identification of phylogenetically conserved regions within the proteins which may play a role in the function of this important enzyme in plant metabolism. Gene specific probes from 3' coding and noncoding regions of the cDNA clone used to probe genomic Southern blots established that this PEPCase gene is present in one copy in the nuclear genome of M. crystallinum. Transcripts arising from this gene increase dramatically when M. crystallinum is irrigated with 0.5 M NaCl, a stress which induces this plant to switch the primary fixation of CO2 from C3 (Calvin cycle) to CAM mode. The salt-induced mRNA encodes a PEPCase isoform which is undetectable in plants in the C3 mode as demonstrated by Northern hybridization.
我们已经确定了编码景天酸代谢（CAM）磷酸烯醇式丙酮酸羧化酶（PEPCase）同基因的全长cDNA的完整核苷酸序列。该cDNA克隆长度为3348 bp，是从经盐胁迫并随后诱导出CAM的冰叶日中花（俗称冰草）中分离的mRNA获得的。这个长开放阅读框编码PEPCase（EC 4.1.1.31），预测分子量为110533道尔顿。冰叶日中花PEPCase推导的氨基酸序列与玉米的最为相似，氨基酸同一性为74.9%。来自大肠杆菌和蓝藻集胞藻的PEPCase蛋白相应区域的序列同一性分别为41.4%和33.5%。这四个氨基酸序列的汇编使得能够鉴定出蛋白质中系统发育保守区域，这些区域可能在植物代谢中这种重要酶的功能中发挥作用。来自cDNA克隆3'编码区和非编码区的基因特异性探针用于探测基因组Southern杂交，结果表明该PEPCase基因在冰叶日中花的核基因组中以单拷贝存在。当用0.5 M NaCl浇灌冰叶日中花时，来自该基因的转录本会急剧增加，这种胁迫会诱导该植物将CO2的初级固定从C3（卡尔文循环）模式转变为CAM模式。如Northern杂交所示，盐诱导的mRNA编码一种在处于C3模式的植物中无法检测到的PEPCase同工型。
