Klemm Clementine, Otto Sebastian, Wolf Constanze, Haseloff Reiner F, Beyermann Michael, Krause Eberhard
Leibniz-Institut für Molekulare Pharmakologie, Robert-Rössle-Str. 10, 13125 Berlin, Germany.
J Mass Spectrom. 2006 Dec;41(12):1623-32. doi: 10.1002/jms.1129.
The affinity of titanium dioxide for phosphate groups has been successfully used for enrichment of phosphopeptides from complex mixtures. This paper reports the relationship between the occurrence of some amino acids and the phospho-specific and nonspecific binding of peptides that occurs during titanium dioxide enrichment. In order to perform a systematic study, two well-characterized peptide mixtures consisting of either 33 or 8 synthetic phosphopeptides and their nonphosphorylated analogs, which differed in charge and hydrophobicity, were synthesized and analyzed by ESI-MS and MALDI-MS. The titanium dioxide procedure was also evaluated for comprehensive detection of phosphopeptides in phosphoproteomics. In summary, our results clearly confirm the high selectivity of titanium dioxide for phosphorylated sequences. Drastically reduced recovery was observed for phosphopeptides with multiple basic amino acids. Nonspecific binding of nonphosphorylated peptides and sample loss of phosphopeptides must also be taken into account.
二氧化钛对磷酸基团的亲和力已成功用于从复杂混合物中富集磷酸化肽段。本文报道了在二氧化钛富集过程中，某些氨基酸的存在与肽段的磷酸化特异性和非特异性结合之间的关系。为了进行系统研究，合成了两种特征明确的肽混合物，分别由33种或8种合成磷酸化肽段及其非磷酸化类似物组成，它们在电荷和疏水性方面存在差异，并通过电喷雾电离质谱（ESI-MS）和基质辅助激光解吸电离质谱（MALDI-MS）进行分析。还评估了二氧化钛方法用于磷酸化蛋白质组学中磷酸化肽段的全面检测。总之，我们的结果清楚地证实了二氧化钛对磷酸化序列具有高选择性。对于含有多个碱性氨基酸的磷酸化肽段，回收率大幅降低。还必须考虑非磷酸化肽段的非特异性结合以及磷酸化肽段的样品损失。
