通过电子图像分析和同源建模解析重组大鼠UBF的结构。

Structure of recombinant rat UBF by electron image analysis and homology modelling.

作者信息

Neil K J, Ridsdale R A, Rutherford B, Taylor L, Larson D E, Glibetic M, Rothblum L I, Harauz G

机构信息

Department of Molecular Biology and Genetics, University of Guelph, Ontario, Canada.

出版信息

Nucleic Acids Res. 1996 Apr 15;24(8):1472-80. doi: 10.1093/nar/24.8.1472.

DOI:10.1093/nar/24.8.1472

原文链接:https://pmc.ncbi.nlm.nih.gov/articles/PMC145827/

Abstract

We have studied the structure of recombinant rat UBF (rrUBF), an RNA polymerase I transcription factor, by electron microscopy and image analysis of single particles contrasted with methylamine tungstate. Recombinant rat UBF appeared to be a flat, U-shaped protein with a central region of low density. In the dominant projections, 2-fold mirror symmetry was seen, consistent with the dimerization properties of this molecule, and of dimensions in agreement with the length of DNA that rat UBF protects in footprinting studies. Electron microscopy of various rrUBF-DNA complexes confirmed that our recombinant protein was fully able to bind the 45S rDNA promoter, and that it caused substantial bends in the DNA. Upon extended incubation in a droplet covered by a lipid monolayer at the liquid-air interface, rrUBF formed long filamentous arrays with a railway track appearance. This structure was interpreted to consist of overlapping rrUBF dimers 3.5 nm apart, which value would represent the thickness of the protein. Our results show rrUBF to interact with and bend the promoter DNA into a roughly 10 nm diameter superhelix. Based on all these electron microscopical results, an atomic structure was predicted by homology modelling of the HMG fingers, and connected by energy minimized intervening segments.

摘要

我们通过电子显微镜和用钨酸甲胺对比的单颗粒图像分析，研究了RNA聚合酶I转录因子重组大鼠UBF（rrUBF）的结构。重组大鼠UBF似乎是一种扁平的U形蛋白质，中央区域密度较低。在主要投影中，观察到二重镜像对称，这与该分子的二聚化特性一致，其尺寸也与大鼠UBF在足迹研究中保护的DNA长度相符。对各种rrUBF-DNA复合物的电子显微镜观察证实，我们的重组蛋白完全能够结合45S rDNA启动子，并使DNA发生明显弯曲。在液-气界面由脂质单层覆盖的液滴中长时间孵育后，rrUBF形成了具有铁轨外观的长丝状阵列。这种结构被解释为由相距3.5 nm的重叠rrUBF二聚体组成，该值代表蛋白质的厚度。我们的结果表明，rrUBF与启动子DNA相互作用，并将其弯曲成直径约10 nm的超螺旋。基于所有这些电子显微镜结果，通过HMG指的同源建模预测了原子结构，并通过能量最小化的中间片段进行连接。

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本文引用的文献

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Structure of the HMG box motif in the B-domain of HMG1.高迁移率族蛋白1（HMG1）B结构域中HMG盒基序的结构

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