Recombinant colorimetric antibodies: construction and characterization of a bifunctional F(ab)2/alkaline phosphatase conjugate produced in Escherichia coli.
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.
Ducancel
F
F
Département d'Ingénierie et d'Etudes des Protéines, C.E.A. Saclay, Gif/Yvette, France.
Gillet
D
D
Carrier
A
A
Lajeunesse
E
E
Ménez
A
A
Boulain
J C
JC
D000469:Alkaline Phosphatase / Q000096:biosynthesis* / Q000235:genetics; D000911:Antibodies, Monoclonal / Q000096:biosynthesis / Q000235:genetics; D001483:Base Sequence; D003124:Colorimetry; D004926:Escherichia coli / Q000235:genetics*; D015870:Gene Expression; D005822:Genetic Vectors; D007140:Immunoglobulin Fab Fragments / Q000096:biosynthesis* / Q000235:genetics; D007074:Immunoglobulin G / Q000737:chemistry / Q000235:genetics; D008969:Molecular Sequence Data; D009876:Operon; D010957:Plasmids; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis* / Q000235:genetics
D000911:Antibodies, Monoclonal; D007140:Immunoglobulin Fab Fragments; D007074:Immunoglobulin G; D011993:Recombinant Fusion Proteins; D000469:Alkaline Phosphatase