Issaq Haleem J, Chan King C, Janini George M, Conrads Thomas P, Veenstra Timothy D
Laboratory of Proteomics and Analytical Technologies, SAIC-Frederick Inc., National Cancer Institute at Frederick, P.O. Box B, Frederick, Maryland, MA 21702, USA.
J Chromatogr B Analyt Technol Biomed Life Sci. 2005 Mar 5;817(1):35-47. doi: 10.1016/j.jchromb.2004.07.042.
Current solution based proteomic analysis methods are generally based on enzymatic digestion of a protein mixture followed by separation using multidimensional liquid chromatography and/or electrophoresis where peptide identification is typically accomplished by tandem mass spectrometry (MS/MS). It is generally accepted that no single chromatographic or electrophoretic procedure is capable of resolving the complex mixture of peptides that results from a global proteolytic digest of a proteome. Therefore, combining two or more orthogonal (multimodal) separation procedures dramatically improves the overall resolution and results in a larger number of peptides being identified from complex proteome digests. Separation of a proteome digest is a particularly challenging analytical problem due to the large number of peptides and the wide concentration dynamic range. While it has been demonstrated that increasing the number of dimensions of separation prior to MS analysis increases the number of peptides that may be identified, a balance between the time invested and the overall results obtained must be carefully considered. This manuscript provides a review of two- and three-dimensional peptide separation strategies combined with MS for the analysis of complex peptide mixtures.
当前基于溶液的蛋白质组学分析方法通常基于对蛋白质混合物进行酶解，然后使用多维液相色谱和/或电泳进行分离，其中肽段鉴定通常通过串联质谱（MS/MS）来完成。人们普遍认为，没有单一的色谱或电泳方法能够分离蛋白质组全局酶解产生的复杂肽段混合物。因此，结合两种或更多种正交（多模式）分离方法可显著提高整体分辨率，并能从复杂蛋白质组酶解物中鉴定出更多的肽段。由于肽段数量众多且浓度动态范围宽广，蛋白质组酶解物的分离是一个极具挑战性的分析问题。虽然已经证明在质谱分析之前增加分离维度的数量会增加可鉴定肽段的数量，但必须仔细权衡投入的时间和获得的整体结果。本文综述了结合质谱用于分析复杂肽段混合物的二维和三维肽段分离策略。
