Dual binding modes for an HMG domain from human HMGB2 on DNA
Dual binding modes for an HMG domain from human HMGB2 on DNA
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DOI：
10.1529/biophysj.104.052068
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发表时间：
2005-07-01
影响因子：
3.4
通讯作者：
Williams, MC
中科院分区：
文献类型：
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作者：
McCauley, M;Hardwidge, PR;Williams, MC
高迁移率族B（HMGB）蛋白含有两个HMG盒结构域，已知其无序列特异性地结合到DNA小沟中，略微插入碱基对之间并在DNA骨架中产生强弯曲。我们使用光学镊子来测量拉伸单个DNA分子所需的力。描述DNA的灵活性，包括轮廓长度和持久性长度的参数，揭示。在存在纳摩尔浓度的从HMGB 2分离的HMG盒A的情况下，DNA显示出其持续长度的减少，其中蛋白质诱导的平均DNA弯曲角对于50 mM Na+为114 +/-21度，对于100 mM Na+为87 +/-9度。DNA轮廓长度从0.341 +/- 0.003增加到0.397 +/- 0.012 nm/碱基对，与盐浓度无关。在50 mM Na+中，蛋白质甚至在高DNA延伸下也不解结合，而在100 mM Na+中，蛋白质似乎仅在低于2 nM的浓度下解结合。这些观察结果支持一个灵活的铰链模型，在低蛋白浓度的非合作HMG结合。然而，在更高的蛋白质浓度，一个合作。观察到的是共振模式而不是铰链结合。这种模式可能是这种强力光镊实验的独特特征。
High mobility group B ( HMGB) proteins contain two HMG box domains known to bind without sequence specificity into the DNA minor groove, slightly intercalating between basepairs and producing a strong bend in the DNA backbone. We use optical tweezers to measure the forces required to stretch single DNA molecules. Parameters describing DNA flexibility, including contour length and persistence length, are revealed. In the presence of nanomolar concentrations of isolated HMG box A from HMGB2, DNA shows a decrease in its persistence length, where the protein induces an average DNA bend angle of 114 +/- 21 degrees for 50 mM Na+, and 87 +/- 9 degrees for 100 mM Na+. The DNA contour length increases from 0.341 +/- 0.003 to 0.397 +/- 0.012 nm per basepair, independent of salt concentration. In 50 mM Na+, the protein does not unbind even at high DNA extension, whereas in 100 mM Na+, the protein appears to unbind only below concentrations of 2 nM. These observations support a flexible hinge model for noncooperative HMG binding at low protein concentrations. However, at higher protein concentrations, a cooperative. lament mode is observed instead of the hinge binding. This mode may be uniquely characterized by this high-force optical tweezers experiment.