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標題: | 鯉魚MKK基因之研究 Studies on Carp Mitogen-Activated Protein Kinase Kinase Genes:Molecular Cloning and Genomic Organizatiion |
作者: | Ming-Shyue Lee 李明學 |
出版年 : | 1994 |
學位: | 碩士 |
摘要: | Mitogen-activated protein kinase kinase(MKK)是一種具有同時對Tyrosine和Serine/Threonine磷酸化的蛋白激脢,調控 Mitogen-activated protein(MAP) kinases的活性。利用RT-PCR在鯉魚卵與肝臟發現兩種MKKs的DNA片段,並進行cDNA基因庫篩選工作,結果只從鯉魚肝臟cDNA基因庫,選殖得到一個含1971bp長度的鯉魚MKK1 cDNA,可轉譯一個含397個氨基酸的蛋白質。並將其轉譯區域的cDNA,在大腸菌中進行基因表現。比較該蛋白質的氨基酸列,發現鯉魚MKK1與rat MKK1, rat MKK2, Xenopus MKK, Drosophila MKK,分別有78%, 80%,76%和58%的氨基酸相同。 另外再以DIG隨機引子法製備探針,進行鯉魚染色體基因庫基因選殖,結果找到鯉魚MKK1和MKK2兩種基因。經核酸定序和基因結構分析,發現鯉魚MKK1基因有11個exons和10個introns,全長7.1kb。最大的intron,長140bp,最小的intron,只有83bp長,但目前仍欠缺啟動子區域序列。 鯉魚MKK2基因,目前只定完部份核酸序列,分析其基因結構,發現該序列包含三個exons和四個introns;其中有一個intron的分佈位置,與鯉魚MKK1第二個intron的位置相同,其餘三個introns的分佈位置,皆不相同。因此,從兩者基因結構比較,鯉魚MKK1和MKK2在演化上可能屬於不同來源的基因。另外,比較氨基酸序列,發現目前已知的鯉魚MKK2與鯉魚MKK1,只有38.2%的氨基酸相同。因此,在魚類或哺乳類動物,可能存在類似酵母菌兩種以上的MAP kinase cascades,以傳遞外界不同的訊息。 We have cloned a full-length cDNA encoding the mitogenactivated protein kinase kinase (MKK) from a carp liver cDNA library. The cDNA contains 1970 bp with a single open reading frame encoding a 397 amino acid protein which is designated as carp MKK1. By comparing with known MKK sequences from other species, carp MKK1 is 78 %, 80 %, 76 % and 58 % identical to rat MKK1, rat MKK2, Xenopus NKK and Drosophila MKK. The structural organization of carp MKK1 gene has also been determined. Carp MKK1 gene comprises eleven exons and ten introns and spans 7.1 Kb. Among those ten introns, the longest is 1.4 Kb whereas the shortest is only 83 bp in length. There is another mkk gene present in carp ovary tissues detected by RT-PCR. However, the work to isolate this kind of cDNA clone from a carp ovary cDNA library is not successful. Only partial sequences of carp mkk2 coding region was obtained by alignment and combination of those sequences from genomic clones and RT-PCR products and these sequences encode 183 amino acid residues. From amino acid sequence comparison, carp MKK2 is 95.6% and 38.2% identical to xenopus MEK2 and carp MKK1, respectively. Therefore, it is possible that there are at least two different MAP kinase cascades involved in the vertebrate signal transduction pathway. |
URI: | http://tdr.lib.ntu.edu.tw/jspui/handle/123456789/76017 |
全文授權: | 未授權 |
顯示於系所單位: | 生化科學研究所 |
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