DETERMINATION OF THE MOLECULAR-WEIGHT OF DNA-BOUND PROTEIN(S) RESPONSIBLE FOR GEL-ELECTROPHORETIC MOBILITY SHIFT OF LINEAR DNA FRAGMENTS EXEMPLIFIED WITH PURIFIED VIRAL MYB PROTEIN
DETERMINATION OF THE MOLECULAR-WEIGHT OF DNA-BOUND PROTEIN(S) RESPONSIBLE FOR GEL-ELECTROPHORETIC MOBILITY SHIFT OF LINEAR DNA FRAGMENTS EXEMPLIFIED WITH PURIFIED VIRAL MYB PROTEIN
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DOI：
10.1093/nar/16.12.5241
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发表时间：
1988-06-24
影响因子：
14.9
通讯作者：
BADING, H
中科院分区：
文献类型：
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作者：
BADING, H
蛋白质-DNA复合物的凝胶电泳迁移率低于游离DNA片段。蛋白质-DNA复合物中线性DNA片段的阻滞程度的一个参数是结合蛋白质的分子量。游离DNA（m）和蛋白质-DNA复合物（m“）的迁移距离的商是结合蛋白质的分子量（MW）的函数。基于在50%非变性聚丙烯酰胺凝胶中对含有lac操纵基因的203 bp DNA片段的lac阻遏物诱导的迁移率变化的评价，可以显示（m/m“-1）与MW之间的正比例关系，比例因子K = 215 kDa。因子K取决于凝胶中丙烯酰胺的浓度，随着丙烯酰胺浓度的增加而降低。计算给出了确定负责的线性DNA片段的电泳迁移率降低的DNA结合因子的分子量。作为一个实例，使用该计算以分析分离的病毒mvb蛋白的DNA结合。可以证明病毒mvb蛋白作为单体和二聚体与DNA结合。
A protein-DNA complex has less gel electrophoretic mobility than the free DNA fragment. One parameter for the degree of retardation of a linear DNA fragment in a protein-DNA complex is the molecular weight of the bound protein(s). The quotient of the migration distances of free DNA (m) and protein-DNA complex (m'') is a function of the molecular weight (MW) of the bound protein(s). Based on the evaluation of the lac repressor induced mobility shift of a 203 bp DNA fragment containing the lac operator in a 50% nondenaturing polyacrylamide gel a direct proportionality could be shown between (m/m''-1) and MW with the proportionality factor K = 215 kDa. The factor K depends on the acrylamide concentration in the gel, getting lower values with increasing acrylamide concentrations. A calculation is given to determine the molecular weight of DNA-binding factors responsible for the decreased electrophoretic mobility of a linear DNA fragment. As an example this calculation was used in order to analyse DNA-binding of the isolated viral mvb protein. It could be demonstrated that the viral mvb protein binds to DNA as a monomer and a dimer.