Gabibov A G, Gololobov G V, Makarevich O I, Schourov D V, Chernova E A, Yadav R P
Engelhardt Institute of Molecular Biology, Russian Academy of Sciences, Moscow.
Appl Biochem Biotechnol. 1994 May-Jun;47(2-3):293-302; discussion 303. doi: 10.1007/BF02787941.
Catalysis by antibodies could be a frequent phenomenon if the immune system generates a sufficiently diverse number of antibody-active sites, some of which may possess catalytic activity. A catalytic antibody can be expected to do more damage than one that simply binds antigen. The best biochemical marker of systemic lupus erythematosus (SLE) is presence of autoantibodies to DNA. In the present article, we describe the DNA-hydrolyzing activity of DNA-binding autoantibodies purified from SLE patients. The substrates employed were supercoiled plasmid, radiolabeled plasmid fragments, and oligonucleotides. Hydrolysis of DNA by the antibodies was indicated by the appearance of fragments visualized by ethidium bromide staining of agarose gels or autoradiography of polyacrylamide gels. Changes in linear dichroism values were also indicative of DNA hydrolysis. The antibody activity was purified by protein A-sepharose chromatography, high-performance liquid chromatography gel filtration, and DNA-affinity chromatography. Scrupulous control studies were done to demonstrate that DNA-hydrolyzing activity really belongs to the antibodies. Purified Fab fragments showed hydrolyzing activity, whereas the Fc fragment was inactive. The specificity of DNA cleavage was investigated, and the rate parameters of hydrolysis by antibodies and conventional nucleases were compared.
如果免疫系统产生足够多样的抗体活性位点，其中一些可能具有催化活性，那么抗体催化可能是一种常见现象。可以预期，催化性抗体造成的损害比单纯结合抗原的抗体更大。系统性红斑狼疮（SLE）的最佳生化标志物是抗DNA自身抗体的存在。在本文中，我们描述了从SLE患者中纯化的DNA结合自身抗体的DNA水解活性。所用的底物是超螺旋质粒、放射性标记的质粒片段和寡核苷酸。通过琼脂糖凝胶溴化乙锭染色或聚丙烯酰胺凝胶放射自显影观察到的片段出现表明抗体对DNA的水解作用。线性二色性值的变化也表明了DNA水解。通过蛋白A-琼脂糖凝胶色谱、高效液相色谱凝胶过滤和DNA亲和色谱纯化抗体活性。进行了严格的对照研究以证明DNA水解活性确实属于抗体。纯化的Fab片段显示水解活性，而Fc片段无活性。研究了DNA切割的特异性，并比较了抗体和传统核酸酶的水解速率参数。
