Protein denaturation has always displayed a huge necessity for mass spectrometry (MS)-based protein identification methods in proteomics. In this research, a novel protein digestion method with the assistance of alternating current (AC) denaturation has been proposed and evaluated. In this method, merely, 200 mM ammonium bicarbonate buffer solution (pH, 8.2) was used to dissolve proteins and act as the electrolyte, and protein denaturation could be achieved in several seconds. For apo-transferrin, ovalbumin and bovine serum albumin that are resistant to digestion in their native states, confident amino acid sequence coverage by matrix assisted laser desorption/ionization-time of flight mass spectrometry (MALDI-TOF MS) analysis were obtained after 200 v AC denaturation. The applicability of this method was further investigated via analyzing a rat liver proteome sample using nano reversed phase liquid chromatography-electrospray ionization-tandem mass spectrometry (nanoRPLC-ESI-MS/MS). As a result, 458 proteins were identified which is comparable to the in-solution digestion via 8 M urea denaturation (375 proteins). All these results demonstrated that AC denaturation could offer an efficient assistance for a clean and high-throughput digestion in the individual level and proteome level.

蛋白质变性始终显示了蛋白质组学中基于质谱（MS）的蛋白质鉴定方法的巨大必要性。在这项研究中，已提出并评估了一种借助交流电（AC）变性的新型蛋白质消化方法。在该方法中，仅使用200 mM碳酸氢铵缓冲溶液（pH，8.2）溶解蛋白质并充当电解质，并且可以在几秒钟内实现蛋白质变性。对于在原始状态下抗消化的脱铁运铁蛋白，卵清蛋白和牛血清白蛋白，在200 v电压下通过基质辅助激光解吸/电离飞行时间质谱（MALDI-TOF MS）分析获得了可靠的氨基酸序列覆盖率AC变性。通过使用纳米反相液相色谱-电喷雾电离串联质谱法（nanoRPLC-ESI-MS / MS）分析大鼠肝脏蛋白质组样品，进一步研究了该方法的适用性。结果，鉴定出了458种蛋白质，与通过8 M尿素变性的溶液内消化相当（375种蛋白质）。所有这些结果表明，AC变性可以为个人水平和蛋白质组水平的清洁和高通量消化提供有效的帮助。