Augustus Anne M, Reardon Patrick N, Spicer Leonard D
Departments of Biochemistry and Radiology, Duke University Medical Center, Durham, NC 27710, USA.
Proc Natl Acad Sci U S A. 2009 Mar 31;106(13):5065-9. doi: 10.1073/pnas.0811130106. Epub 2009 Mar 16.
Atomic level characterization of proteins and other macromolecules in the living cell is challenging. Recent advances in NMR instrumentation and methods, however, have enabled in-cell studies with prospects for multidimensional spectral characterization of individual macromolecular components. We present NMR data on the in-cell behavior of the MetJ repressor from Escherichia coli, a protein that regulates the expression of genes involved in methionine biosynthesis. NMR studies of whole cells along with corresponding studies in cell lysates and in vitro preparations of the pure protein give clear evidence for extensive nonspecific interactions with genomic DNA. These interactions can provide an efficient mechanism for searching out target sequences by reducing the dependence on 3-dimensional diffusion through the crowded cellular environment. DNA provides the track for MetJ to negotiate the obstacles inherent in cells and facilitates locating and binding specific repression sites, allowing for timely control of methionine biosynthesis.
对活细胞中蛋白质和其他大分子进行原子水平的表征具有挑战性。然而,核磁共振(NMR)仪器和方法的最新进展使得在细胞内进行研究成为可能,有望对单个大分子成分进行多维光谱表征。我们展示了来自大肠杆菌的MetJ阻遏蛋白在细胞内行为的核磁共振数据,该蛋白调节参与蛋氨酸生物合成的基因表达。对全细胞的核磁共振研究以及在细胞裂解物和纯蛋白的体外制剂中的相应研究,为与基因组DNA的广泛非特异性相互作用提供了明确证据。这些相互作用可以通过减少在拥挤的细胞环境中对三维扩散的依赖,提供一种寻找靶序列的有效机制。DNA为MetJ提供了在细胞固有障碍中穿行的轨道,并有助于定位和结合特定的阻遏位点,从而及时控制蛋氨酸的生物合成。