Applying Arginylation for Bottom-Up Proteomics.
Arginylation is an enzymatic reaction in which arginyl-tRNA protein transferase 1 (ATE1, EC 2.3.2.8) conjugates a single arginyl moiety from aminoacylated tRNA(Arg) onto a target polypeptide. We established arginylation for in vitro labeling of peptides with N-terminal acidic amino acids. Consistent with prior knowledge, arginylated peptides flanked by basic amino acids result in rich redundant MS/MS fragment spectra using various precursor fragmentation modes. Arginylation carried out by ATE1 is a fast method for labeling peptides. Sequence-specific proteolytic digest of proteins is best carried out using a double digest of proteins by Lys-C and Asp-N to generate peptides with a basic amino acid on the C-terminus and an acidic amino acid on the N-terminus. Under these conditions, arginylation is specific for N-terminal acidic amino acids and results in a near 2× sequence coverage in the MS/MS spectrum are achieved.
Institute of Molecular Systems Biology, Eidgenössische Technische Hochschule (ETH) Zürich, HPT E 61, Auguste-Piccard-Hof 1, 8093, Zürich, Switzerland, ebhardt@imsb.biol.ethz.ch.
H Alexander
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Ebhardt
D019881:Aminoacyltransferases / Q000378:metabolism*; D001120:Arginine / Q000378:metabolism*; D013058:Mass Spectrometry / Q000379:methods; D010455:Peptides / Q000378:metabolism; D011499:Protein Processing, Post-Translational; D011506:Proteins / Q000378:metabolism*; D040901:Proteomics / Q000379:methods; D013379:Substrate Specificity
D010455:Peptides; D011506:Proteins; D001120:Arginine; D019881:Aminoacyltransferases; C019727:arginyltransferase