Ducancel F, Gillet D, Carrier A, Lajeunesse E, Ménez A, Boulain J C
Département d'Ingénierie et d'Etudes des Protéines, C.E.A. Saclay, Gif/Yvette, France.
Biotechnology (N Y). 1993 May;11(5):601-5. doi: 10.1038/nbt0593-601.
We have designed a vector which allows the synthesis in Escherichia coli of bifunctional F(ab)2-alkaline phosphatase conjugates. The vector contains a di-cistronic operon encoding truncated heavy chain (Fd or VH-CH1) of an IgG inserted between residues +6 and +7 of bacterial alkaline phosphatase (PhoA), and the light chain of the same IgG. We demonstrate the utility of this approach with the heavy and light chain domains of a snake toxin-specific monoclonal antibody, M alpha 2-3. We show that the VH-CH1-PhoA hybrid and VL-CL are concomitantly expressed and exported to the periplasm of E. coli where they form a disulfide-linked chimeric protein. The hybrid has the same affinity as M alpha 2-3 for the snake toxin antigen and possesses PhoA enzymatic activity.
我们设计了一种载体，可在大肠杆菌中合成双功能F(ab)2-碱性磷酸酶偶联物。该载体包含一个双顺反子操纵子，编码插入到细菌碱性磷酸酶（PhoA）第+6和+7位残基之间的IgG截短重链（Fd或VH-CH1）以及同一IgG的轻链。我们用蛇毒素特异性单克隆抗体Mα2-3的重链和轻链结构域证明了这种方法的实用性。我们表明，VH-CH1-PhoA杂种和VL-CL伴随表达并输出到大肠杆菌周质中，在那里它们形成二硫键连接的嵌合蛋白。该杂种对蛇毒素抗原具有与Mα2-3相同的亲和力，并具有PhoA酶活性。
