Lys-C/Arg-C, a More Specific and Efficient Digestion Approach for Proteomics Studies

Abstract

Nowadays, bottom-up approaches are predominantly adopted in proteomics studies, which necessitate a proteolysis step prior to MS analysis. Trypsin is often the best protease in choice due to its high specificity and MS-favored proteolytic products. A lot of efforts have been made to develop a superior digestion approach but hardly succeed, especially in large-scale proteomics studies. Herein, we report a new tandem digestion using Lys-C and Arg-C, termed Lys-C/Arg-C, which has been proven to be more specific and efficient than trypsin digestion. Reanalysis of our previous data ( Anal. Chem. 2018, 90 (3), 1554-1559) revealed that both Lys-C and Arg-C are trypsin-like proteases and perform better when considered as trypsin. In particular, for Arg-C, the identification capacity is increased to 2.6 times and even comparable with trypsin. The good complementarity, high digestion efficiency, and high specificity of Lys-C and Arg-C prompt the Lys-C/Arg-C digestion. We systematically evaluated Lys-C/Arg-C digestion using qualitative and quantitative proteomics approaches and confirmed its superior performance in digestion specificity, efficiency, and identification capacity to the currently widely used trypsin and Lys-C/trypsin digestions. As a result, we concluded that the Lys-C/Arg-C digestion approach would be the choice of next-generation digestion approach in both qualitative and quantitative proteomics studies. Data are available via ProteomeXchange with identifier PXD009797.