Foettinger Alexandra, Leitner Alexander, Lindner Wolfgang
Department of Analytical Chemistry and Food Chemistry, University of Vienna, Waehringer Strasse 38, 1090 Vienna, Austria.
J Proteome Res. 2007 Sep;6(9):3827-34. doi: 10.1021/pr0702767. Epub 2007 Jul 27.
A method for the selective enrichment of tryptophan-containing peptides from complex peptide mixtures such as protein digests is presented. It is based on the reversible reaction of tryptophan with malondialdehyde and trapping of the derivatized Trp-peptides on hydrazide beads via the free aldehyde group of the modified peptides. The peptides are subsequently recovered in their native form by specific cleavage reactions for further (mass spectrometric) analysis. The method was optimized and evaluated using a tryptic digest of a mixture of 10 model proteins, demonstrating a significant reduction in sample complexity while still allowing the identification of all proteins. The applicability of the tryptophan-specific enrichment procedure to complex biological samples is demonstrated for a total yeast cell lysate. Analysis of the processed fraction by 1D-LC-MS/MS confirms the specificity of the enrichment procedure, as more than 85% of the peptides recovered from the enrichment step contained tryptophan. The reduction in sample complexity also resulted in the identification of additional proteins in comparison to the untreated lysate.
本文介绍了一种从复杂肽混合物（如蛋白质消化产物）中选择性富集含色氨酸肽的方法。该方法基于色氨酸与丙二醛的可逆反应，以及通过修饰肽的游离醛基将衍生化的色氨酸肽捕获在酰肼珠上。随后，通过特异性裂解反应以天然形式回收肽，用于进一步（质谱）分析。使用10种模型蛋白质混合物的胰蛋白酶消化产物对该方法进行了优化和评估，结果表明样品复杂性显著降低，同时仍能鉴定所有蛋白质。对于全酵母细胞裂解物，证明了色氨酸特异性富集程序对复杂生物样品的适用性。通过一维液相色谱-串联质谱对处理后的部分进行分析，证实了富集程序的特异性，因为从富集步骤回收的肽中超过85%含有色氨酸。与未处理的裂解物相比，样品复杂性的降低还导致鉴定出了更多蛋白质。
