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Applying Arginylation for Bottom-Up Proteomics

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Protein Arginylation

Part of the book series: Methods in Molecular Biology ((MIMB,volume 1337))

Abstract

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 tRNAArg 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.

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Acknowledgments

This work was supported by a FP7 Marie Curie International Incoming Fellowship to HAE.

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Authors and Affiliations

  1. Institute of Molecular Systems Biology, Eidgenössische Technische Hochschule (ETH) Zürich, HPT E 61, Auguste-Piccard-Hof 1, 8093, Zürich, Switzerland

    H. Alexander Ebhardt

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  1. H. Alexander Ebhardt
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Correspondence to H. Alexander Ebhardt .

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Editors and Affiliations

  1. Department of Animal Biology, University of Pennsylvania School of Veterinary Medicine, Philadelphia, Pennsylvania, USA

    Anna S. Kashina

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Ebhardt, H.A. (2015). Applying Arginylation for Bottom-Up Proteomics. In: Kashina, A. (eds) Protein Arginylation. Methods in Molecular Biology, vol 1337. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-2935-1_16

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  • DOI: https://doi.org/10.1007/978-1-4939-2935-1_16

  • Publisher Name: Humana Press, New York, NY

  • Print ISBN: 978-1-4939-2934-4

  • Online ISBN: 978-1-4939-2935-1

  • eBook Packages: Springer Protocols

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