Fregeau D R, Davis P A, Danner D J, Ansari A, Coppel R L, Dickson E R, Gershwin M E
Division of Rheumatology, Allergy and Clinical Immunology, University of California, Davis 95616.
J Immunol. 1989 Jun 1;142(11):3815-20.
Antimitochondrial antibodies (AMA) recognizing the acetyltransferase (E2) of the pyruvate dehydrogenase (PDH) complex have been previously well-documented and the immunodominant epitope mapped. In this study, we demonstrate that sera from patients with primary biliary cirrhosis (PBC) react with another lipoic acid containing acyltransferase enzyme, namely the E2 of the branched chain alpha-ketoacid dehydrogenase (BCKD) complex. Indeed, 85/120 (71%) sera from patients with PBC reacted with BCKD-E2 by immunoblotting against purified BCKD complex. In contrast, sera from patients with chronic active hepatitis or progressive sclerosing cholangitis as well as sera from healthy volunteers did not react with any component enzymes of the BCKD complex. More importantly, BCKD enzyme activity was inhibited after incubation of the BCKD complex with either PBC sera against BCKD-E2 or with affinity purified antisera to BCKD-E2. Enzyme activity was unaltered by control sera or with PBC sera that reacted with PDH-E2 but not BCKD-E2. Furthermore, immunoblots of purified mitochondria probed with PBC sera absorbed with BCKD-E2 demonstrated that BCKD-E2 and PDH-E2 are each recognized by distinct AMA populations which do not cross-react. In addition, affinity purified PBC sera against BCKD-E2 did not react with PDH-E2 nor inhibit PDH enzyme activity, thus providing further evidence that BCKD-E2 and PDH-E2 are recognized by separate AMA. These data further suggest that the BCKD-E2 epitope recognized by AMA contains, or is close to, a functional domain of this enzyme. The availability of cDNA clones encoding BCKD-E2 and PDH-E2 will allow the study of how key metabolic enzymes may be involved in the immunology and pathology of PBC.
识别丙酮酸脱氢酶（PDH）复合体乙酰转移酶（E2）的抗线粒体抗体（AMA）此前已有充分文献记载，其免疫显性表位也已定位。在本研究中，我们证明原发性胆汁性肝硬化（PBC）患者的血清可与另一种含硫辛酸的酰基转移酶发生反应，即支链α-酮酸脱氢酶（BCKD）复合体的E2。实际上，通过针对纯化的BCKD复合体进行免疫印迹分析，120例PBC患者中有85例（71%）的血清与BCKD-E2发生反应。相比之下，慢性活动性肝炎或进行性硬化性胆管炎患者的血清以及健康志愿者的血清均未与BCKD复合体的任何组成酶发生反应。更重要的是，将BCKD复合体与针对BCKD-E2的PBC血清或与亲和纯化的抗BCKD-E2血清孵育后，BCKD酶活性受到抑制。对照血清或与PDH-E2而非BCKD-E2发生反应的PBC血清不会改变酶活性。此外，用与BCKD-E2吸附的PBC血清检测纯化线粒体的免疫印迹表明，BCKD-E2和PDH-E2各自被不同的AMA群体识别，且不发生交叉反应。此外，亲和纯化的针对BCKD-E2的PBC血清不与PDH-E2反应，也不抑制PDH酶活性，从而进一步证明BCKD-E2和PDH-E2被不同的AMA识别。这些数据进一步表明，AMA识别的BCKD-E2表位包含该酶的一个功能域或与之接近。编码BCKD-E2和PDH-E2的cDNA克隆的可得性将有助于研究关键代谢酶如何参与PBC的免疫学和病理学过程。
