頁籤選單縮合
題名 | Characterization, Cloning, and Expression of Porcine αB Crystallin= |
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作者 | Liao,Jiahn-haur; Hung,Chin-chun; Lee,Jiahn-shing; Wu,Shih-hsiung; Chiou,Shyh-horng; |
期刊 | 中央研究院生物化學研究所論文集 |
出版日期 | 19980000 |
卷期 | 24 1998[民87.] |
頁次 | 頁75-81 |
分類號 | 361.19 |
語文 | eng |
關鍵詞 | αB crystallin; α-Crystallin; |
英文摘要 | α-Crystallin is a major lens protein present in the lenses of all vertebrate species. Recent studies have revealed that bovine a-crystallins possess genuine chaperone activity similar to small heat-shock proteins. In order to compare this chaperone-like structural protein from the eye lenses of different mammalian species, we have cloned and expressed one of the main α-crystallin subunits, i.e., α B crystallin, from the porcine lenses in order to facilitate the structure-function evaluation and comparison of this chaperonin protein. cDNA encoding αB subunit chain was obtained using a new "Marathon cDNA amplification" protocol of Polymerase Chain Reaction (PCR). PCR-amplified product corresponding to αB subunit was then ligated into pGEM-T plasmid and prepared for nucleotide sequencing by the dideoxynucleotide chain-termination method. Sequencing several positive clones containing DNA inserts coding for αB-crystallin subunit constructed only one complete full-length reading frame of 525 base pairs similar to human and bovine αB subunits, covering a deduced protein sequence of 175 amino acids including the universal translation-initiating methionine. The porcine αB crystallin shows only 3 and 7 residues difference to bovine and human αB crystallins respectively, revealing the close relatedness among mammalian eye lens proteins. The sequence differences between porcine and submammalian species such as chicken and bullfrog are much greater, especially at the N- and C-terminal regions of these αB crystallins. Expression of αB subunit chain in E. coli vector generated a polypeptide which can cross-react with the antiserum against the native and purified αB subunit from the native porcine lenses albeit with a much lower activity. |
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