Epitope mapping of monoclonal antibodies against 4-aminobenzoate hydroxylase from Agaricus bisporus.
Four monoclonal antibodies (mAbs) against 4-aminobenzoate hydroxylase (EC 1.14.13.27) have been produced (H. Tsuji et al., J. Biol. Chem. 265 (1990) 16064; T. Ogawa et al., Biochim. Biophys. Acta 1115 (1992) 220). Of the mAbs, three mAbs (mAb-A, -B1 and -B2) recognize the FAD-binding domain of the enzyme. In the present study, the epitopes of the mAbs on the enzyme have been examined using pGEX-2T expression systems for DNA fragments encoding various partial amino acid sequences of 4-aminobenzoate hydroxylase. The epitopes for mAb-A, -B1 and -B2 were shown to be on sequences 413-434, 435-460 and 380-413, respectively. These findings suggest that these epitopes for mAb-A, -B1 and -B2 may be close to the isoalloxazine moiety of FAD, which plays a central role in the catalysis of the enzyme.
Tsuji
H
H
Department of Nutritional Science, Faculty of health and Welfare Science, Okayama Prefectural University, Soja, Okayama 719-1197, Japan.
Kimoto
M
M
Watanabe
H
H
Sasagawa
T
T
Oka
T
T
Yamashita
H
H
Okita
M
M
D000364:Agaricus / Q000201:enzymology*; D000911:Antibodies, Monoclonal / Q000737:chemistry / Q000276:immunology*; D004247:DNA / Q000096:biosynthesis; D000939:Epitopes / Q000032:analysis*; D004926:Escherichia coli / Q000378:metabolism; D005182:Flavin-Adenine Dinucleotide / Q000276:immunology; D005982:Glutathione Transferase / Q000737:chemistry / Q000276:immunology; D006899:Mixed Function Oxygenases / Q000737:chemistry / Q000276:immunology*; D010957:Plasmids; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis / Q000737:chemistry / Q000276:immunology*
D000911:Antibodies, Monoclonal; D000939:Epitopes; D011993:Recombinant Fusion Proteins; D005182:Flavin-Adenine Dinucleotide; D004247:DNA; D006899:Mixed Function Oxygenases; C050092:4-aminobenzoate hydroxylase; D005982:Glutathione Transferase