染色质亚基引发针对核蛋白抗原决定簇的物种特异性抗体。

Tahourdin C S, Bustin M

Biochemistry. 1980 Sep 16;19(19):4387-94. doi: 10.1021/bi00560a002.

Nucleosomes composed of 195 base pairs of DNA associated with histones H2A, H2B, H3, and H4 purified from chicken erythrocyte nuclei were used to elicit antibodies in rabbits. Specific serological reaction between the antisera and the nucleosomes is demonstrated by immunodiffusion, immunofluorescence, microcomplement fixation, solid-phase radioimmunoassay, immunosedimentation, and polyacrylamide gel electrophoresis of 5'-32P end-labeled nucleosomes. The antisera did not react with DNA extracted from these nucleosomes, core histones, or the cross-linked histone octamer from chicken erythocytes, calf thymus total histones, or chromosomal proteins HMG-1 or HMG-17. Nucleosome antigenicity was not affected by redigestion with micrococcal nuclease. Digestion with DNase I brought about 50% loss of reactivity while digestion with trypsin or proteinase K resulted in total loss of activity. The antisera reacted strongly with trimer, dimer, and monomer nucleosomes as well as with the core particle (145 base pairs of DNA) and subnucleosome (greater than 145 base pairs) obtained from chicken. It reacted less well with nucleosomes obtained from HeLa cells and was almost totally devoid of activity against chromatin particles obtained from rat liver or wheat germ. Experiments employing the technique of transferring proteins from a polyacrylamide gel to diazobenzyloxymethyl paper and visualization of antigens by autoradiography excluded the possibility that the serum contains antibodies against tissue-specific antigens which are found in small amounts but are very immunogenic. It is concluded that most of the anitbodies in the sera are directed against nucleoprotein antigenic determinants composed of the N-terminal portion of the histones and segments of DNA. Antibody binding is dependent on contact between the histone and DNA segments and is independent of the integrity of the entire nucleosome. Thus, certain histone DNA contacts remain intact even though the structure of the nucleosome has been disrupted.

由从鸡红细胞核中纯化得到的与组蛋白H2A、H2B、H3和H4结合的195个碱基对的DNA组成的核小体，被用于在兔体内引发抗体。抗血清与核小体之间的特异性血清学反应通过免疫扩散、免疫荧光、微量补体结合、固相放射免疫测定、免疫沉淀以及对5'-32P末端标记核小体的聚丙烯酰胺凝胶电泳来证明。抗血清不与从这些核小体中提取的DNA、核心组蛋白、鸡红细胞的交联组蛋白八聚体、小牛胸腺总组蛋白或染色体蛋白HMG-1或HMG-17发生反应。核小体的抗原性不受微球菌核酸酶再消化的影响。用DNA酶I消化导致反应性丧失50%，而用胰蛋白酶或蛋白酶K消化导致活性完全丧失。抗血清与三聚体、二聚体和单体核小体以及从鸡获得的核心颗粒（145个碱基对的DNA）和亚核小体（大于145个碱基对）强烈反应。它与从HeLa细胞获得的核小体反应较差，并且几乎完全没有针对从大鼠肝脏或小麦胚芽获得的染色质颗粒的活性。采用将蛋白质从聚丙烯酰胺凝胶转移到重氮苄氧基甲基纸上并通过放射自显影观察抗原的技术进行的实验排除了血清中含有针对组织特异性抗原的抗体的可能性，这些组织特异性抗原含量少但免疫原性强。得出的结论是，血清中的大多数抗体针对由组蛋白的N末端部分和DNA片段组成的核蛋白抗原决定簇。抗体结合依赖于组蛋白和DNA片段之间的接触，并且独立于整个核小体的完整性。因此，即使核小体的结构已被破坏，某些组蛋白-DNA接触仍保持完整。