Hill John J, Laue Thomas M
Amgen, Seattle, Washington, USA.
Center to Advance Molecular Interaction Science, University of New Hampshire, Durham, New Hampshire, USA.
Methods Enzymol. 2015;562:501-27. doi: 10.1016/bs.mie.2015.06.012. Epub 2015 Jul 14.
This chapter illustrates how analytical ultracentrifugation methods, coupled with the fluorescence detection system, are an excellent approach to characterizing and comparing protein-binding interactions in dilute solution and concentrated, crowded solutions like serum. We show that in serum, the binding and assembly states for a pair of endogenous protein ligands and an antibody inhibitor are dramatically different than those observed in dilute, simple buffers. This type of analysis approach may be helpful in research efforts intent at discerning the underpinnings to a therapeutic's activity and pharmacokinetic properties in vivo.
本章阐述了分析超速离心方法与荧光检测系统相结合,是表征和比较稀溶液以及血清等浓缩、拥挤溶液中蛋白质结合相互作用的绝佳方法。我们表明,在血清中,一对内源性蛋白质配体和一种抗体抑制剂的结合及组装状态与在稀的简单缓冲液中观察到的情况显著不同。这种分析方法可能有助于旨在洞察治疗药物在体内的活性和药代动力学特性基础的研究工作。