Pfitzner R, Wagener R, Stoffel W
Biol Chem Hoppe Seyler. 1986 Oct;367(10):1077-83. doi: 10.1515/bchm3.1986.367.2.1077.
The isolation and characterization of a human apolipoprotein B 100-specific cDNA clone (lambda gt-B1) containing a 1321 base pairs (bp) spanning insert is described. It encodes the 3'-nontranslated 281 bp long region up to the polyadenylation site and 1040 bp of the C-terminal coding region of 345 amino-acid residues of human apo B 100 and the stop codon. The lambda gt-B1 cDNA clone has been isolated from a human hepatoma cDNA expression library by immunoscreening using affinity-purified polyclonal anti apo B 100 antibodies. The nucleotide sequence of the apo B 100 insert has been determined. A part of the polypeptide sequence derived from this nucleotide sequence was identical with the amino-acid sequence obtained by protein sequencing of a purified cyanogen bromide fragment of apo B 100. The fusion protein consisting of beta-galactosidase and the 345 amino-acid residue long C-terminus of apo B 100 had an apparent molecular mass of 148 kDa in NaDodSO4 polyacrylamide gel electrophoresis. In Northern blot hybridization analysis the insert of the apo B 100-cDNA clone hybridized to a 20 to 22 kb mRNA from adult human liver.
本文描述了一个人载脂蛋白B 100特异性cDNA克隆（λgt - B1）的分离和特性鉴定，该克隆包含一个1321个碱基对（bp）的插入片段。它编码直至聚腺苷酸化位点的3'非翻译区281 bp长区域、人载脂蛋白B 100 345个氨基酸残基的C末端编码区1040 bp以及终止密码子。λgt - B1 cDNA克隆是通过使用亲和纯化的多克隆抗载脂蛋白B 100抗体进行免疫筛选，从人肝癌cDNA表达文库中分离得到的。已确定载脂蛋白B 100插入片段的核苷酸序列。从该核苷酸序列推导的部分多肽序列与通过对纯化的载脂蛋白B 100溴化氰片段进行蛋白质测序获得的氨基酸序列相同。在十二烷基硫酸钠聚丙烯酰胺凝胶电泳中，由β - 半乳糖苷酶和载脂蛋白B 100的345个氨基酸残基长的C末端组成的融合蛋白的表观分子量为148 kDa。在Northern印迹杂交分析中，载脂蛋白B 100 - cDNA克隆的插入片段与来自成人肝脏的20至22 kb mRNA杂交。
