Giuliani M M, Ricci S, Ratti G, Pucci P, Marino G, Malorni A, Ceccarini C, Terrana B, Tecce M F
Cell and Molecular Biology Laboratories, Sclavo Research Center, Siena, Italy.
Protein Eng. 1989 Aug;2(8):605-10. doi: 10.1093/protein/2.8.605.
A DNA sequence coding for human alpha-fetoprotein amino acid sequence 38-119 was synthesized and cloned in a bacterial expression vector. The alpha-fetoprotein sequence was selected as the least homologous to albumin, since the two proteins have an overall amino acid identity of approximately 38%. A chimeric protein was obtained which was purified by preparative electrophoresis and characterized in its primary structure by fast atom bombardment mass spectometry. About 70% of the alpha-fetoprotein sequence was physically mapped and found to correspond to the amino acids encoded in the synthetic gene. The use of this recombinant protein allowed the selection of monoclonal antibodies recognizing both the recombinant fragment and native alpha-fetoprotein. These antibodies should allow the development of an immunoassay for alpha-fetoprotein with absolute selectivity versus albumin. This might result in more sensitive clinical determinations, avoiding the possibility of cross-reactions.
合成了编码人甲胎蛋白氨基酸序列38 - 119的DNA序列，并将其克隆到细菌表达载体中。选择甲胎蛋白序列与白蛋白同源性最低的部分，因为这两种蛋白质的总体氨基酸同一性约为38%。获得了一种嵌合蛋白，通过制备电泳进行纯化，并通过快原子轰击质谱法对其一级结构进行了表征。约70%的甲胎蛋白序列被进行了物理定位，发现其与合成基因中编码的氨基酸相对应。使用这种重组蛋白使得能够筛选出既能识别重组片段又能识别天然甲胎蛋白的单克隆抗体。这些抗体应能开发出一种针对甲胎蛋白的免疫测定法，对白蛋白具有绝对选择性。这可能会带来更灵敏的临床检测结果，避免交叉反应的可能性。
