Cloning and expression of a murine Fab fragment recognizing a defined linear epitope of Chironomus thummi thummi major allergen Chi t 1-9.
We have cloned and expressed in bacteria the genes coding for the Fab fragment of the monoclonal antibody (MoAb) M3. M3 is a murine IgG1 antibody reactive with the major allergen Chi t 1-9 of Chironomus thummi thummi. The major allergen Chi t 1-9 is known to be an aggressive inhalant allergen and causes type-I allergy. The immunoglobulin (Ig) fragment genes were cloned as a synthetic dicistronic operon. In this operon each gene is preceded by a bacterial signal sequence to direct the recombinant protein to the periplasmic space of the bacteria. The cloned genes were expressed in Escherichia coli using the strong T7-RNA-polymerase-based system. Sequence analysis revealed that the light and heavy chains of MoAb M3 belong to the V kappa II and V kappa IIC group of the Ig family, respectively. Genes of the V kappa II and V kappa IIC group are known to be used in response to haptens. The apparent affinity constants of the parent antibody M3 and the recombinant Fab fragment are nearly equivalent.
Kipp
B
B
Borstel Research Institute, Division of Allergy, Germany.
Schlaak
M
M
Becker
W M
WM
D000485:Allergens / Q000276:immunology*; D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D002683:Chironomidae / Q000276:immunology*; D003001:Cloning, Molecular; D000939:Epitopes / Q000276:immunology*; D005803:Genes, Immunoglobulin; D006454:Hemoglobins / Q000276:immunology*; D007140:Immunoglobulin Fab Fragments / Q000737:chemistry* / Q000235:genetics* / Q000302:isolation & purification; D019476:Insect Proteins; D051379:Mice; D008969:Molecular Sequence Data
D000485:Allergens; C071697:Chi t I allergen; D000939:Epitopes; D006454:Hemoglobins; D007140:Immunoglobulin Fab Fragments; D019476:Insect Proteins