title: Differential isotope dansylation labeling combined with liquid chromatography mass spectrometry for quantification of intact and N-terminal truncated proteins.
abstract: The N-terminal amino acids of proteins are important structure units for maintaining the biological function, localization, and interaction networks of proteins. Under different biological conditions, one or several N-terminal amino acids could be cleaved from an intact protein due to processes, such as proteolysis, resulting in the change of protein properties. Thus, the ability to quantify the N-terminal truncated forms of proteins is of great importance, particularly in the area of development and production of protein-based drugs where the relative quantity of the intact protein and its truncated form needs to be monitored. In this work, we describe a rapid method for absolute quantification of protein mixtures containing intact and N-terminal truncated proteins. This method is based on dansylation labeling of the N-terminal amino acids of proteins, followed by microwave-assisted acid hydrolysis of the proteins into amino acids. It is shown that dansyl labeled amino acids are stable in acidic conditions and can be quantified by liquid chromatography mass spectrometry (LC-MS) with the use of isotope analog standards.
authors.affiliation: Department of Chemistry, University of Alberta, Edmonton, Alberta, Canada.
authors.forename: Yanan
authors.initials: Y
authors.lastname: Tang
authors.forename: Liang
authors.initials: L
authors.lastname: Li
mesh_terms: D002478:Cells, Cultured; D002623:Chemistry Techniques, Analytical / Q000379:methods*; D002853:Chromatography, Liquid; D003619:Dansyl Compounds / Q000032:analysis*; D004926:Escherichia coli; D007553:Isotope Labeling; D057230:Limit of Detection; D013058:Mass Spectrometry; D011506:Proteins / Q000032:analysis*; D013997:Time Factors
chemical_list: D003619:Dansyl Compounds; D011506:Proteins; C033853:dansyl acid
keywords: Dansylation; Isotope labeling; Liquid chromatography; Mass spectrometry; Microwave-assisted acid hydrolysis; Protein quantification