Separation and structural analysis of IgG-binding minor proteins in human plasma

我们研究了人血浆微量蛋白的来源，我们发现在两种不同的尺寸分离条件下（二维电泳），我们比较了二维凝胶电泳的模式；一个在整个运行过程中没有变性剂，一个在大小分离电泳过程中存在SDS。利用基质辅助激光解吸电离飞行时间质谱（MALDI-TOF MS）对蛋白质进行肽段质量指纹图谱鉴定，利用电喷雾电离串联质谱（ESI-MS/MS）对部分氨基酸进行质量标签法测序。此外，我们开发了一种三步电泳技术，在等电聚焦的步骤中使用琼脂糖凝胶，并表明这对次要蛋白质的结构分析是有用的。通过比较利用不同条件的双向电泳得到的图谱，以及对图谱上蛋白质的结构分析，将有助于了解功能性蛋白质复合物的构建过程。

We have examined the origin of human plasma minor proteins which we found comparing the patterns of two-dimensional gel electrophoresis run under two different conditions of size-separation (second dimension electrophoresis) ; one in the absence of denaturing agents throughout the run and one in the presence of SDS during the size-separation electrophoresis. The minor proteins were analyzed by mass spectrometry, identification of proteins by peptide mass fingerprinting using matrix-assisted laser desorption ionization-time-of-flignt mass spectrometer (MALDI-TOF MS) and partial amino acid sequencing by mass-tag method using electrospray ionization tandem mass spectrometer (ESI-MS/MS). Further, we have developed a three-step electrophoresis technique which employ slab-agarose gels in the step of isoelectric focusing and showed that this is useful for the structural analysis of the minor proteins. The comparisons of the patterns obtained by using different conditions of two-dimensional electrophoresis, together with the structural analyses of the proteins on the patterns, will provide insights on the construction processes of functional protein complexes.