Experimental test of connector rotation during DNA packaging into bacteriophage φ29 capsids
Experimental test of connector rotation during DNA packaging into bacteriophage φ29 capsids
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DOI：
10.1371/journal.pbio.0050059
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发表时间：
2007-03-01
期刊：
影响因子：
9.8
通讯作者：
Bustamante, Carlos
中科院分区：
文献类型：
--
作者：
Hugel, Thorsten;Michaelis, Jens;Bustamante, Carlos
噬菌体phi 29产生巨大的力，通过门静脉马达复合体将其双链DNA基因组压缩成蛋白质衣壳。几个机械模型的产生这些高的力量，由电机复合预测耦合的DNA易位旋转的头-尾连接器十二聚体。假定的连接器旋转在这里研究相结合的方法，单分子力谱偏振敏感单分子荧光。在我们的实验中，我们观察电机功能在几个包装复合物中使用视频显微镜的珠位置在磁阱平行。同时，我们遵循连接到门静脉马达连接器的单个荧光团的方向。根据我们的数据，我们可以排除连接器旋转的可能性大于99%，因此回答了一个长期存在的机械问题。
The bacteriophage phi 29 generates large forces to compact its double-stranded DNA genome into a protein capsid by means of a portal motor complex. Several mechanical models for the generation of these high forces by the motor complex predict coupling of DNA translocation to rotation of the head-tail connector dodecamer. Putative connector rotation is investigated here by combining the methods of single-molecule force spectroscopy with polarizationsensitive single-molecule fluorescence. In our experiment, we observe motor function in several packaging complexes in parallel using video microscopy of bead position in a magnetic trap. At the same time, we follow the orientation of single fluorophores attached to the portal motor connector. From our data, we can exclude connector rotation with greater than 99% probability and therefore answer a long-standing mechanistic question.