Abstract

Proteolytic digestion prior to LC–MS analysis is a key step for the identification of proteins. Digestion of proteins is typically performed with trypsin, but certain proteins or important protein sequence regions might be missed using this endoproteinase. Only few alternative endoproteinases are available and chemical cleavage of proteins is rarely used. Recently, it has been reported that some metal complexes can act as artificial proteases. In particular, the Lewis acid scandium(III) triflate has been shown to catalyze the cleavage of peptide bonds to serine and threonine residues. Therefore, we investigated if this compound can also be used for the cleavage of proteins. For this purpose, several single proteins, the 20S immune-proteasome (17 proteins), and the Universal Proteomics Standard UPS1 (48 proteins) were analyzed by MALDI–MS and/or LC–MS. A high cleavage specificity N-terminal to serine and threonine residues was observed, but also additional peptides with deviating cleavage specificity were found. Scandium(III) triflate can be a useful tool in protein analysis as no other reagent has been reported yet which showed cleavage specificity within proteins to serines and threonines.

Graphic abstract

中文翻译：

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使用三氟甲磺酸scan主要切割丝氨酸和苏氨酸N末端的蛋白质

摘要

LC-MS分析之前的蛋白水解消化是鉴定蛋白质的关键步骤。蛋白质的消化通常使用胰蛋白酶进行，但是使用此内切蛋白酶可能会遗漏某些蛋白质或重要的蛋白质序列区域。仅有几种可供选择的内蛋白酶可用，并且很少使用蛋白质的化学裂解。最近，已经报道了一些金属络合物可以充当人工蛋白酶。特别是，路易斯酸三氟甲磺酸scan（III）已显示出催化肽键裂解为丝氨酸和苏氨酸残基的能力。因此，我们研究了该化合物是否也可用于蛋白质切割。为此目的，几种单一蛋白，即20S免疫蛋白酶体（17种蛋白），MALDI-MS和/或LC-MS分析了通用蛋白质组学标准UPS1（48种蛋白质）。观察到对丝氨酸和苏氨酸残基N-末端的高切割特异性，但是还发现了具有不同切割特异性的其他肽。三氟甲磺酸（（III）在蛋白质分析中可能是有用的工具，因为尚未有其他试剂显示出蛋白质对丝氨酸和苏氨酸的切割特异性。

图形摘要