Studies on the Role of Choline/Ethanolamine Kinase in Cell Growth.

利用多肽序列信息，从大鼠肾脏cDNA文库中分离到多个编码42kDa的胆碱/乙醇胺激酶（CK/EK）的cDNA克隆。对8个克隆进行了测序，所有克隆都得到了相同的重叠核苷酸序列。其中4个具有完整的开放阅读框，可编码394个氨基酸，计算分子大小为45100。预测的氨基酸序列包含纯化的42kDa酶的所有内部肽片段序列。当将开放阅读框引入pGEX-2T载体并转染到大肠杆菌细胞时，细胞裂解液中确实出现了显著的CK/EK活性。与先前报道的大鼠肝脏CK cDNA同源性比较显示，全核苷酸序列为66-68%，氨基酸序列为57-59%，表明本文克隆的cDNA一定是新的CK/EK基因产物。从交配后17天的小鼠胚胎cDNA文库中分离出与大鼠42kDa CK/EK和50kDa CK同源的互补dna，并对其序列进行比较。小鼠与大鼠之间50kDa CK/EKs的核苷酸序列同源性（编码序列内）为96.0%，显著高于小鼠与大鼠之间42kDa CK/EKs的同源性（92.4%）。对几种大鼠组织的Northern blot和RT-PCR研究表明，这两种同工酶均普遍表达，且在睾丸中表达水平最高。

By means of peptide sequence information, several cDNA clones encoding a 42kDa choline/ethanolamine kinase (CK/EK) were isolated from a rat kidney cDNA library. Eight clones were sequenced with all of them resulting in identical overlapping nucleotide sequences. Four of them posessed entire open reading frame which could encode 394 amino acids with a calculated molecular size of 45,100. The predicted amino acid sequence contained all of the internal petide fragment sequences derived from the purified 42kDa enzyme. When the open reading frame was introduced into pGEX-2T vector and transfected into E.coli cells, a significant CK/EK activity did appear in the cell lysate. A homology comparison with the previously reported CK cDNAs from rat liver showed 66-68% in entire nucleotide sequences and 57-59% in amino acid sequences, indicating that the cloned cDNA here must be a novel CK/EK gene product.Complementary DNAs homologous to a rat 42kDa CK/EK and to a 50kDa CK were isolated from a 17-day post coitum mouse embryo cDNA library and their sequences were compared btween the two murine species, respectively. The nucleotide sequence homology (within the coding sequence) between mouse and rat 50kDa CKs (96.0%) was considerably higher than that between their 42kDa CK/EKs (92.4%). Northern blot and RT-PCR studies on several rat tissues demonstrated that both isozymes are expressed ubiquitously with the highest level in testis.