Satoh H, Martin B M, Schulick A H, Christ D D, Kenna J G, Pohl L R
Laboratory of Chemical Pharmacology, National Heart, Lung and Blood Institute, Bethesda, MD 20892.
Proc Natl Acad Sci U S A. 1989 Jan;86(1):322-6. doi: 10.1073/pnas.86.1.322.
Previous studies have demonstrated that patients with halothane-induced hepatitis have serum antibodies that are directed against novel liver microsomal neoantigens and have suggested that these neoantigens may play an immunopathological role in development of the patients' liver damage. These investigations have further revealed that the antibodies are directed against distinct polypeptide fractions (100 kDa, 76 kDa, 59 kDa, 57 kDa, 54 kDa) that have been covalently modified by the reactive trifluoroacetyl halide metabolite of halothane. In this paper, the trifluoroacetylated (TFA) 59-kDa neoantigen (59-kDa-TFA) recognized by the patients' antibodies was isolated from liver microsomes of halothane-treated rats by chromatography on an immunoaffinity column of anti-TFA IgG. Antibodies were raised against the 59-kDa-TFA protein and were used to purify the native protein from liver microsomes of untreated rats. Based upon its apparent monomeric molecular mass, NH2-terminal amino acid sequence, catalytic activity, and other physical properties, the protein has been identified as a previously characterized microsomal carboxylesterase (EC 3.1.1.1). A similar strategy may be used to purify and characterized neoantigens associated with other drug toxicities that are believed to have an immunopathological basis.
先前的研究表明,氟烷诱导的肝炎患者血清中存在针对新型肝微粒体新抗原的抗体,并提示这些新抗原可能在患者肝损伤的发生中起免疫病理作用。这些研究进一步揭示,这些抗体针对的是已被氟烷的活性三氟乙酰卤代谢产物共价修饰的不同多肽组分(100 kDa、76 kDa、59 kDa、57 kDa、54 kDa)。在本文中,通过在抗三氟乙酰化IgG免疫亲和柱上进行色谱分离,从氟烷处理大鼠的肝微粒体中分离出患者抗体识别的三氟乙酰化(TFA)59-kDa新抗原(59-kDa-TFA)。制备了针对59-kDa-TFA蛋白的抗体,并用于从未经处理大鼠的肝微粒体中纯化天然蛋白。根据其明显的单体分子量、氨基末端氨基酸序列、催化活性和其他物理性质,该蛋白已被鉴定为先前已表征的微粒体羧酸酯酶(EC 3.1.1.1)。类似的策略可用于纯化和表征与其他被认为具有免疫病理基础的药物毒性相关的新抗原。