Ramisetty Sravani K, Langlete Petter, Lale Rahmi, Dias Rita S
Biophysics and Medical Technology, Department of Physics, NTNU Norwegian University of Science and Technology, NO-7491 Trondheim, Norway.
Department of Biotechnology, PhotoSynLab, Faculty of Natural Sciences and Technology, NTNU Norwegian University of Science and Technology, NO-7491 Trondheim, Norway.
Int J Biol Macromol. 2017 Oct;103:845-853. doi: 10.1016/j.ijbiomac.2017.05.079. Epub 2017 May 20.
The macromolecules of the bacterial cell occupy 20-40% of the total cytosol volume, and crowded environments have long been known to compact and stabilize DNA. Nevertheless, investigations on DNA-protein binding are generally performed in the absence of crowding, which may yield an incomplete understanding of how nucleoid-assembling proteins work. A family of such proteins, abundant in Gram-negative bacteria, is the histone-like nucleoid structuring proteins (H-NS). Herein, the synergistic role of macromolecular crowding (mimicked using polyethylene glycol, PEG) and H-NS was investigated using fluorescence correlation spectroscopy (FCS) and enzyme protection assays. We show that crowding enhances the binding of H-NS to the AT-rich tracks of the DNA, where it preferentially binds to, protecting these tracks towards enzyme digestion, inducing some DNA condensation, and inhibiting the biological function of DNA. We further suggest that the looping of DNA chains, induced by H-NS, contributes to the synergistic effect of DNA-binding protein and crowding agents, on DNA condensation.
细菌细胞中的大分子占据了细胞质总体积的20%-40%，而人们早就知道拥挤的环境会使DNA压缩并稳定。然而，关于DNA-蛋白质结合的研究通常是在不存在拥挤的情况下进行的，这可能导致对类核组装蛋白的工作方式理解不完整。一类在革兰氏阴性菌中大量存在的此类蛋白质是类组蛋白类核结构蛋白（H-NS）。在此，使用荧光相关光谱法（FCS）和酶保护试验研究了大分子拥挤（用聚乙二醇，PEG模拟）和H-NS的协同作用。我们表明，拥挤增强了H-NS与DNA富含AT的序列的结合，H-NS优先结合于此，保护这些序列免受酶消化，诱导一些DNA凝聚，并抑制DNA的生物学功能。我们进一步表明，由H-NS诱导的DNA链环化有助于DNA结合蛋白和拥挤剂对DNA凝聚的协同作用。
