Physikalische und Theoretische Chemie - NanoBioSciences, Technische Universität Braunschweig, Hans-Sommer-Strasse 10, 38106 Braunschweig, Germany.
Curr Opin Chem Biol. 2013 Aug;17(4):691-8. doi: 10.1016/j.cbpa.2013.05.020. Epub 2013 Jun 13.
Fluorescence spectroscopy and fluorescence microscopy carried out on the single molecule level are elegant methods to decipher complex biological systems; it can provide a wealth of information that frequently is obscured in the averaging of ensemble measurements. Fluorescence can be used to localise a molecule, study its binding with interaction partners and ligands, or to follow conformational changes in large multicomponent systems. Efficient labelling of proteins and nucleic acids is very important for any fluorescence method, and equally the development of novel fluorophores has been crucial in making biomolecules amenable to single molecule fluorescence methods. In this paper we review novel coupling strategies that permit site-specific and efficient labelling of proteins. Furthermore, we will discuss progressive single molecule approaches that allow the detection of individual molecules and biomolecular complexes even directly isolated from cellular extracts at much higher and much lower concentrations than has been possible so far.
荧光光谱学和荧光显微镜技术在单分子水平上的应用是解析复杂生物系统的优雅方法；它可以提供大量信息，而这些信息通常在对整体测量结果进行平均时会被掩盖。荧光可用于定位分子、研究其与相互作用伙伴和配体的结合情况，或跟踪大的多组分系统中的构象变化。对任何荧光方法来说，蛋白质和核酸的有效标记都非常重要，而新型荧光团的开发对于使生物分子能够适应单分子荧光方法也是至关重要的。在本文中，我们将综述允许蛋白质进行定点和高效标记的新型偶联策略。此外，我们还将讨论渐进式单分子方法，这些方法允许在比以往更高和更低的浓度下，直接从细胞提取物中检测到单个分子和生物分子复合物。
