N-ethylmaleimide inhibition of the DNA-binding activity of the herpes simplex virus type 1 major DNA-binding protein.
N-ethylmaleimide inhibition of the DNA-binding activity of the herpes simplex virus type 1 major DNA-binding protein.
复制标题
N-乙基马来酰亚胺抑制单纯疱疹病毒 1 型主要 DNA 结合蛋白的 DNA 结合活性。
DOI：
10.1128/jvi.62.3.810-817.1988
复制
发表时间：
1988
影响因子：
5.4
通讯作者：
Ruyechan,WT
中科院分区：
文献类型：
--
作者：
Ruyechan,WT
主要的单纯疱疹病毒DNA结合蛋白，命名为ICP 8，与单链DNA紧密结合，是病毒DNA复制所必需的。的DNA结合活性的ICP 8的巯基试剂N-乙基马来酰亚胺的行动的敏感性已被检查通过使用硝酸纤维素过滤结合和琼脂糖凝胶电泳分析。ICP 8与N-乙基马来酰亚胺的孵育导致DNA结合活性的快速丧失。ICP 8与单链DNA的预孵育显著抑制这种结合活性的丧失。这些结果意味着游离巯基参与了ICP 8与单链DNA的相互作用，并且该巯基在结合后变得不太容易接近环境。在存在和不存在N-乙基马来酰亚胺的情况下的结合相互作用的琼脂糖凝胶电泳分析表明，ICP 8所表现出的合作结合在用该试剂处理后丢失，但可能仍然存在一些残留的非合作结合。最后一个结果通过用32 P标记的寡核苷酸dT 10和天然和N-乙基马来酰亚胺处理的ICP 8进行的平衡透析实验得到证实。
The major herpes simplex virus DNA-binding protein, designated ICP8, binds tightly to single-stranded DNA and is required for replication of viral DNA. The sensitivity of the DNA-binding activity of ICP8 to the action of the sulfhydryl reagent N-ethylmaleimide has been examined by using nitrocellulose filter-binding and agarose gel electrophoresis assays. Incubation of ICP8 with N-ethylmaleimide results in a rapid loss of DNA-binding activity. Preincubation of ICP8 with single-stranded DNA markedly inhibits this loss of binding activity. These results imply that a free sulfhydryl group is involved in the interaction of ICP8 with single-stranded DNA and that this sulfhydryl group becomes less accessible to the environment upon binding. Agarose gel electrophoretic analysis of the binding interaction in the presence and absence of N-ethylmaleimide indicates that the cooperative binding exhibited by ICP8 is lost upon treatment with this reagent but that some residual noncooperative binding may remain. This last result was confirmed by equilibrium dialysis experiments with the 32P-labeled oligonucleotide dT10 and native and N-ethylmaleimide-treated ICP8.