Gerlich W H, Robinson W S
Cell. 1980 Oct;21(3):801-9. doi: 10.1016/0092-8674(80)90443-2.
Hepatitis B virus DNA contains a tightly bound protein which was not removed by healing to 60 degrees C with 2% SDS, 2% mercaptoethanol. The protein was indirectly demonstrated by the extraction of the DNA-protein complex with phenol before but not after its digestion with proteinase K. The DNA-protein complex had a lower buoyant density than protease-treated or free DNA; it was bound to glass fiber filters; it migrated at a slower rate in gel electrophoresis; and it could be radiolabeled by oxidative iodination. The binding site of the protein was mapped by extraction of restriction endonuclease digests with phenol and analysis of the digests for missing DNA fragments. The protein was localized to a site near the 5' end of the complete viral DNA strand. It remained attached to this strand after heating with SDS to 90 degrees C or treatment with 0.1 N NaOH, suggesting a covalent linkage. The 5' end of neither viral DNA strand could be phosphorylated in a reaction with polynucleotide kinase, consistent with attachment of protein to the 5' ends. The incomplete DNA strand, however, which is the strand elongated by the virion DNA polymerase reaction, did not contain a detectable amount of polypeptide as did the complete strand. The reasons for the apparent block of the 5' end of the incomplete DNA strand is thus not known. The protein bound covalently to HBV DNA could be involved in the replication of the complete viral DNA strand and/or endonucleolytic generation of linear unit-length DNA pieces from replicative intermediates, although its function and origin are not yet known.
乙肝病毒DNA含有一种紧密结合的蛋白质，用2%十二烷基硫酸钠（SDS）和2%巯基乙醇在60℃处理并不能将其去除。在用蛋白酶K消化之前，通过苯酚提取DNA - 蛋白质复合物间接证明了该蛋白质的存在，消化后则无法证明。DNA - 蛋白质复合物的浮力密度低于蛋白酶处理过的或游离的DNA；它能与玻璃纤维滤膜结合；在凝胶电泳中迁移速度较慢；并且可以通过氧化碘化进行放射性标记。通过用苯酚提取限制性内切酶消化产物并分析消化产物中缺失的DNA片段来定位该蛋白质的结合位点。该蛋白质定位于完整病毒DNA链5'端附近的一个位点。在用SDS加热至90℃或用0.1N氢氧化钠处理后，它仍附着于该链上，提示存在共价连接。在与多核苷酸激酶的反应中，病毒DNA两条链的5'端均不能被磷酸化，这与蛋白质附着于5'端一致。然而，不完全DNA链，即通过病毒体DNA聚合酶反应延长的链，与完整链不同，它不含可检测量的多肽。因此，不完全DNA链5'端明显受阻的原因尚不清楚。与乙肝病毒DNA共价结合的蛋白质可能参与完整病毒DNA链的复制和/或从复制中间体进行的线性单位长度DNA片段的内切核酸酶生成，尽管其功能和来源尚不清楚。
