Shikhman A R, Greenspan N S, Cunningham M W
Department of Microbiology and Immunology, University of Oklahoma Health Science Center, Oklahoma City 73190.
J Immunol. 1993 Oct 1;151(7):3902-13.
It is well known that antibodies to N-acetyl-beta-D-glucosamine (GlcNAc) cross-react with cardiac valves, skin, and other host tissues. However, molecular targets of these antibodies have not been identified. For this reason, anti-streptococcal mAb cross-reactive with group A streptococci and heart proteins were studied for their reactivity with GlcNAc, the immunodominant epitope of group A streptococcal carbohydrate. Characterization of the mAb that recognized GlcNAc revealed that each mAb had its own unique antigen-binding profile and pattern of immunofluorescence on rat heart cells. In the ELISA and Western blot these mAb reacted with cytoskeletal and heart proteins such as actin, keratin, myosin, and vimentin, as well as with streptococcal recombinant M5 and M6 proteins. Binding of the mAb to cytoskeletal proteins was inhibited by GlcNAc conjugated with BSA in a dose-dependent manner, and the mAb preferentially reacted with high-density GlcNAc-BSA conjugates. Antigenic determinants on the proteins recognized by the mAb were resistant to sodium periodate and N-acetylglucosaminidase treatment, suggesting reactivity with peptide and not carbohydrate structures. On reaction of the mAb with a panel of synthetic streptococcal, viral, and myosin peptides, one of the mAb, 49.8.9, was found to react most strongly with a synthetic peptide sequence synthesized from the coxsackievirus B3 capsid protein VP1, which shows homology with and cross-reacts with sequences in the streptococcal M6 protein and human cardiac myosin. This most interesting mAb, previously shown to neutralize coxsackie viruses, recognized the amino acid sequence RRKLEFF, which may mimic the GlcNAc epitope. The data collected show that we have identified a new group of multireactive autoantibodies that recognize GlcNAc and cytoskeletal proteins, as well as defined peptide epitopes.
众所周知，抗N-乙酰-β-D-葡萄糖胺（GlcNAc）抗体与心脏瓣膜、皮肤及其他宿主组织发生交叉反应。然而，这些抗体的分子靶点尚未明确。因此，研究了与A组链球菌和心脏蛋白交叉反应的抗链球菌单克隆抗体与A组链球菌碳水化合物免疫显性表位GlcNAc的反应性。对识别GlcNAc的单克隆抗体的特性分析表明，每种单克隆抗体都有其独特的抗原结合谱和在大鼠心脏细胞上的免疫荧光模式。在酶联免疫吸附测定（ELISA）和蛋白质印迹法中，这些单克隆抗体与细胞骨架蛋白和心脏蛋白如肌动蛋白、角蛋白、肌球蛋白和波形蛋白反应，也与链球菌重组M5和M6蛋白反应。与牛血清白蛋白（BSA）偶联的GlcNAc以剂量依赖方式抑制单克隆抗体与细胞骨架蛋白的结合，且单克隆抗体优先与高密度GlcNAc-BSA偶联物反应。单克隆抗体识别的蛋白质上的抗原决定簇对高碘酸钠和N-乙酰葡糖胺酶处理具有抗性，表明其与肽结构而非碳水化合物结构发生反应。在用一组合成的链球菌、病毒和肌球蛋白肽与单克隆抗体反应时，发现其中一种单克隆抗体49.8.9与由柯萨奇病毒B3衣壳蛋白VP1合成的合成肽序列反应最强，该序列与链球菌M6蛋白和人心脏肌球蛋白中的序列具有同源性并发生交叉反应。这种最有趣的单克隆抗体先前已被证明可中和柯萨奇病毒，它识别氨基酸序列RRKLEFF，该序列可能模拟GlcNAc表位。收集的数据表明，我们已经鉴定出一组新的多反应性自身抗体，它们识别GlcNAc、细胞骨架蛋白以及特定的肽表位。
