Cysteine-reactive covalent capture tags for enrichment of cysteine-containing peptides.
Considering the tremendous complexity and the wide dynamic range of protein samples from biological origin and their proteolytic peptide mixtures, proteomics largely requires simplification strategies. One common approach to reduce sample complexity is to target a particular amino acid in proteins or peptides, such as cysteine (Cys), with chemical tags in order to reduce the analysis to a subset of the whole proteome. The present work describes the synthesis and the use of two new cysteinyl tags, so-called cysteine-reactive covalent capture tags (C3T), for the isolation of Cys-containing peptides. These bifunctional molecules were specifically designed to react with cysteines through iodoacetyl and acryloyl moieties and permit efficient selection of the tagged peptides. To do so, a thioproline was chosen as the isolating group to form, after a deprotection/activation step, a thiazolidine with an aldehyde resin by the covalent capture (CC) method. The applicability of the enrichment strategy was demonstrated on small synthetic peptides as well as on peptides derived from digested proteins. Mass spectrometric (MS) analysis and tandem mass spectrometric (MS/MS) sequencing confirmed the efficient and straightforward selection of the cysteine-containing peptides. The combination of C3T and CC methods provides an effective alternative to reduce sample complexity and access low abundance proteins.
Giron
Priscille
P
Biomedical Proteomics Group, Department of Structural Biology and Bioinformatics, University of Geneva, CH-1211 Geneva 4, Switzerland.
Dayon
Loïc
L
Mihala
Nikolett
N
Sanchez
Jean-Charles
JC
Rose
Keith
K
D000818:Animals; D003432:Cross-Linking Reagents / Q000737:chemistry*; D003545:Cysteine / Q000032:analysis / Q000737:chemistry*; D006736:Horses; D006801:Humans; D013058:Mass Spectrometry / Q000379:methods*; D009211:Myoglobin / Q000032:analysis / Q000737:chemistry; D010446:Peptide Fragments / Q000032:analysis / Q000737:chemistry*; D011506:Proteins / Q000032:analysis / Q000737:chemistry*; D012709:Serum Albumin / Q000032:analysis / Q000737:chemistry; D053778:Thiazolidines / Q000737:chemistry
D003432:Cross-Linking Reagents; D009211:Myoglobin; D010446:Peptide Fragments; D011506:Proteins; D012709:Serum Albumin; D053778:Thiazolidines; D003545:Cysteine