Hydrophobic residues 382-386 of antithrombin III, Ala-Ala-Ala-Ser-Thr, serve as the epitope for an antibody which facilitates hydrolysis of the inhibitor by thrombin.
In the presence of a monoclonal antibody raised against the human thrombin-antithrombin III complex, the reaction between thrombin and antithrombin III proceeded to form preferentially a two-chain form of the inhibitor rather than to follow the major pathway of stable acyl complex formation. We thus propose the term "switching antibody" for an antibody that switches the enzyme-inhibitor reaction (Asakura, S., Matsuda, M., Yoshida, N., Terukina, S., and Kihara, H. (1989) J. Biol. Chem. 264, 13736-13739). By analyzing a CNBr fragment of the thrombin-antithrombin III complex that reacts with the antibody we localized the epitope for the antibody to a strongly hydrophobic residue 382-386 peptide segment, Ala-Ala-Ala-Ser-Thr, of the inhibitor, which is also contiguous with a hydrophobic amino acid Ala at its carboxyl terminus. This particular region should be cryptic in nascent antithrombin III, but could have been exposed to provide the reactive site for the antibody at an early stage of the reaction. Thereby a conformational change may have been induced at or near the reactive site of the complex, facilitating hydrolysis of the inhibitor by the enzyme. Interestingly, this hydrophobic region is highly conserved among members of the serpin family.
Division of Hemostasis and Thrombosis Research, Jichi Medical School, Tochigi-Ken, Japan.
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Asakura
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Hirata
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Okazaki
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Hashimoto-Gotoh
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Matsuda
D000595:Amino Acid Sequence; D000911:Antibodies, Monoclonal; D000990:Antithrombin III / Q000276:immunology / Q000378:metabolism*; D003488:Cyanogen Bromide; D000939:Epitopes / Q000276:immunology*; D006801:Humans; D006868:Hydrolysis; D015151:Immunoblotting; D007700:Kinetics; D008961:Models, Structural; D008969:Molecular Sequence Data; D009842:Oligopeptides / Q000494:pharmacology; D010446:Peptide Fragments / Q000302:isolation & purification; D011487:Protein Conformation; D013917:Thrombin / Q000378:metabolism*
D000911:Antibodies, Monoclonal; D000939:Epitopes; D009842:Oligopeptides; D010446:Peptide Fragments; D000990:Antithrombin III; D013917:Thrombin; D003488:Cyanogen Bromide