Yu Ying-Qing, Gilar Martin, Lee Peter J, Bouvier Edouard S P, Gebler John C
Life Sciences Research and Development, Waters Corporation, 34 Maple Street, Milford, Massachusetts 01757, USA.
Anal Chem. 2003 Nov 1;75(21):6023-8. doi: 10.1021/ac0346196.
Improved in-solution tryptic digestion of proteins in terms of speed and peptide coverage was achieved with the aid of a novel acid-labile anionic surfactant (ALS). Unlike SDS, ALS solubilizes proteins without inhibiting trypsin or other common endopeptidases activity. Trypsin activity was evaluated in the presence of various denaturants; little or no decrease in proteolytic activity was observed in 0.1-1% ALS solutions (w/v). Sample preparation prior to mass spectrometry and liquid chromatography analysis consists of sample acidification. ALS degrades rapidly at low-pH conditions, which eliminates surfactant-caused interference with analysis. Described methodology combines the advantages of protein solubilization, rapid digestion, high peptide coverages, and easy sample preparation for mass spectrometry and liquid chromatography analyses.
借助一种新型酸不稳定阴离子表面活性剂(ALS),在蛋白质的溶液内胰蛋白酶消化方面,速度和肽段覆盖度都得到了提高。与十二烷基硫酸钠(SDS)不同,ALS可溶解蛋白质,而不会抑制胰蛋白酶或其他常见内肽酶的活性。在各种变性剂存在的情况下评估了胰蛋白酶的活性;在0.1 - 1% ALS溶液(w/v)中,未观察到蛋白水解活性有显著降低或几乎没有降低。质谱分析和液相色谱分析之前的样品制备包括样品酸化。ALS在低pH条件下会迅速降解,从而消除了表面活性剂对分析的干扰。所描述的方法结合了蛋白质溶解、快速消化、高肽段覆盖度以及易于进行质谱分析和液相色谱分析的样品制备等优点。