Enantioselective immunorecognition of protein modification with optically active ibuprofen using polyclonal antibody.
Formation of covalently bound protein adducts with 2-arylpropionic acids (2-APAs) has been proposed as a possible explanation for hypersensitivity and toxic responses to chiral carboxylic acid drugs. To identify the cellular proteins chemically modified with optically active (S)-ibuprofen, we generate polyclonal antibodies by immunizing rabbits with immunogen coupled to bovine serum albumin (BSA) via the spacer of 4-aminobutyric acid. The resulting antibodies largely cross-reacted with N-alpha-(t-butoxycarbonyl)--(S)-ibuprofenyl lysine as well as with the conjuguated (S)-ibuprofen with glycine and taurine and unconjugated (S)-ibuprofen, enabling enantioselective detection of (S)-ibuprofen residues anchored on ovalbumin molecules, introduced by the reaction of the ibuprofen p-nitrophenyl ester. Furthermore, immunoblotting with an antibody allows the enantioselective detection of (S)-ibuprofen-introduced glutathione-S-transferase (GST). These results indicate that the developed method will be useful for monitoring the generation and localization of protein covalently bound with (S)-ibuprofen, which may be the cause of ibuprofen-induced toxicity.
Ito
Hiromi
H
Faculty of Pharmaceutical Sciences, Kinki University, 3-4-1 Kowakae, Higashi-osaka 577-8502, Japan.
Ishiwata
Shunji
S
Kosaka
Takeshi
T
Nakashima
Rika
R
Takeshita
Harunori
H
Negoro
Sakiko
S
Maeda
Masako
M
Ikegawa
Shigeo
S
D000595:Amino Acid Sequence; D000906:Antibodies / Q000276:immunology*; D004591:Electrophoresis, Polyacrylamide Gel; D007052:Ibuprofen / Q000737:chemistry* / Q000276:immunology; D009682:Magnetic Resonance Spectroscopy; D008969:Molecular Sequence Data; D011506:Proteins / Q000737:chemistry*; D013237:Stereoisomerism
D000906:Antibodies; D011506:Proteins; D007052:Ibuprofen