Bioengineering Graduate Program, University of Kansas, Lawrence, Kansas 66045, USA.
J Pharm Sci. 2013 Jan;102(1):43-51. doi: 10.1002/jps.23367. Epub 2012 Nov 15.
A solution to the problem of being able to show statistically significant differences in the measurements of various levels of higher-order protein structure has been an elusive one. We propose the use of comparative signature diagrams (CSDs) to this end. CSDs compare datasets from different biophysical techniques that fingerprint the secondary, tertiary, and quaternary structures of a protein molecule and display statistically significant differences in these datasets. In this paper, we explore the differences in the structures of two proteins (Granulocyte Colony Stimulating Factor [GCSF] and a monoclonal antibody [mAb]) in various formulations. These proteins were chosen based on the extent of differences in structure observed in the formulations. As an initial test, we utilize data from circular dichroism, 8-anilino-1-naphthalene-sulfonate and intrinsic fluorescence spectroscopy, and static light scattering measurements to fingerprint protein structure in the different formulations. Several layers of statistics were explored to visualize the regions of significant differences in the protein spectra. This approach provides a rapid, high-resolution methodology to compare various structural levels of proteins using standard biophysical instrumentation.
能够在各种高级蛋白质结构的测量中显示出统计学上显著差异的问题一直难以解决。为此，我们提出使用比较特征图谱（CSD）的方法。CSD 比较来自不同生物物理技术的数据，这些技术可以记录蛋白质分子的二级、三级和四级结构，并显示这些数据集中的统计学显著差异。在本文中，我们探讨了两种蛋白质（粒细胞集落刺激因子[GCSF]和单克隆抗体[mAb]）在不同配方中的结构差异。选择这些蛋白质是基于在配方中观察到的结构差异程度。作为初步测试，我们利用圆二色性、8-苯胺-1-萘磺酸和内源荧光光谱以及静态光散射测量的数据来鉴定不同配方中蛋白质的结构。探索了几个统计层次来可视化蛋白质光谱中显著差异的区域。这种方法提供了一种快速、高分辨率的方法，使用标准的生物物理仪器比较蛋白质的各种结构层次。
