Shimizu M, Miyake M, Kanke F, Matsumoto U, Shindo H
Laboratory of Analytical Chemistry, School of Pharmacy, Tokyo University of Pharmacy and Life Science, Japan.
Biochim Biophys Acta. 1995 Dec 27;1264(3):330-6. doi: 10.1016/0167-4781(95)00173-5.
The binding of E. coli histone-like protein HU to curved and uncurved DNA fragments containing adenine tracts was characterized by relative binding affinity assay, and compared with that of other homologous histone-like protein integration host factor (IHF). Both HU and IHF have about 3- to 5-fold higher affinity for overall curved DNA fragments such as (A6N4)11 and (A3T3N4)12 compared to a standard duplex fragment with mixed sequence. The binding manner of HU to the curved fragments was highly cooperative. However, loss of overall curvature for shorter fragments (< approximately 100 bp) reduced the preference of HU binding to curved (A3T3N4)n over uncurved (T3A3N4)n, indicating that the binding specificity of HU to curved DNA is length-dependent. Thus, the curved DNA configuration of the whole molecule facilitates the binding of several HU molecules to form the hierarchy of HU-DNA complex. Furthermore, it was shown that HU and IHF bind less well to (A6N9)n, which has a zig-zag straight structure, whereas they preferentially bind to uncurved (T3A3N4)14. These results suggested that not only intrinsically overall curvature but also the preferred orientations for DNA bending in the protein-DNA complex are important factors for affinities of HU and IHF.
通过相对结合亲和力测定对大肠杆菌类组蛋白HU与含有腺嘌呤序列的弯曲和非弯曲DNA片段的结合进行了表征，并与其他同源类组蛋白整合宿主因子（IHF）进行了比较。与具有混合序列的标准双链体片段相比，HU和IHF对整体弯曲的DNA片段（如（A6N4）11和（A3T3N4）12）的亲和力都高约3至5倍。HU与弯曲片段的结合方式具有高度协同性。然而，较短片段（<约100 bp）整体曲率的丧失降低了HU对弯曲的（A3T3N4）n比对非弯曲的（T3A3N4）n的结合偏好，这表明HU对弯曲DNA的结合特异性是长度依赖性的。因此，整个分子的弯曲DNA构型促进了几个HU分子的结合，以形成HU-DNA复合物的层次结构。此外，研究表明，HU和IHF与具有锯齿状直结构的（A6N9）n结合较差，而它们优先结合非弯曲的（T3A3N4）14。这些结果表明，不仅内在的整体曲率，而且蛋白质-DNA复合物中DNA弯曲的优选方向都是影响HU和IHF亲和力的重要因素。
