Abstract
Multiple Reaction Monitoring Initiated Detection and Sequencing (MIDAS™) is a mass spectrometry-based technique for the detection and characterization of specific post-translational modifications (Unwin et al. 4:1134–1144, 2005), for example acetylated lysine residues (Griffiths et al. 18:1423–1428, 2007). The MIDAS™ technique has application for discovery and analysis of acetylation sites. It is a hypothesis-driven approach that requires a priori knowledge of the primary sequence of the target protein and a proteolytic digest of this protein. MIDAS essentially performs a targeted search for the presence of modified, for example acetylated, peptides. The detection is based on the combination of the predicted molecular weight (measured as mass–charge ratio) of the acetylated proteolytic peptide and a diagnostic fragment (product ion of m/z 126.1), which is generated by specific fragmentation of acetylated peptides during collision induced dissociation performed in tandem mass spectrometry (MS) analysis. Sequence information is subsequently obtained which enables acetylation site assignment. The technique of MIDAS was later trademarked by ABSciex for targeted protein analysis where an MRM scan is combined with full MS/MS product ion scan to enable sequence confirmation.
This is a preview of subscription content, log in via an institution to check access.
Access this chapter
Tax calculation will be finalised at checkout
Purchases are for personal use only
References
Yang XJ, Seto E (2008) Lysine acetylation: codified cross talk with other post translational modifications. Mol Cell 31:449–461
Walther TC, Mann M (2010) Mass spectrometry-based proteomics in cell biology. J Cell Biol 190:491–500
Mischerikow N, Heck AJ (2011) Targeted large-scale analysis of protein acetylation. Proteomics 11:571–589
Griffiths J, Unwin R, Evans C, Leech S, Corfe B, Whetton A (2007) The application of hypothesis driven strategy to the sensitive detection and location of acetylated lysine residues. JASMS 18:1423–1428
Leech SH, Evans CA, Shaw L, Wong CH, Connolly J, Griffiths JR, Whetton AD, Corfe BM (2008) Proteomic analyses of intermediate filaments reveals cytokeratin 8 is highly acetylated—implications for colorectal epithelial homeostasis. Proteomics 8:279–288
Drake PJ, Grififths GJ, Shaw L, Benson RP, Corfe BM (2009) Application of high-content analysis to the study of post-translational modifications of the cytoskeleton. J Proteome Res 8:28–34
Borchers C, Parker CE, Deterding LJ, Tomer KB (1999) Preliminary comparison of precursor scans and liquid chromatography-tandem mass spectrometry on a hybrid quadrupole time-of-flight mass spectrometer. J Chromatogr A 854:119–130
Kim JY, Kim KW, Kwon HJ, Lee DW, Yoo JS (2002) Probing lysine acetylation with a modification specific marker ion using high-performance liquid chromatography/electrospray-mass spectrometry with collision-induced dissociation. Anal Chem 74:5443–5449
Dormeyer W, Ott M, Schnolzer M (2005) Probing lysine acetylation in proteins: strategies, limitations, and pitfalls of in vitro acetyltransferase assays. Mol Cell Proteomics 4:1226–1239
Unwin RD, Griffiths JR, Whetton AD (2009) A sensitive mass spectrometric method for hypothesis-driven detection of peptide post-translational modifications: multiple reaction monitoring-initiated detection and sequencing (MIDAS). Nat Protoc 4:870–877
Perkins DN, Pappin DJ, Creasy DM, Cottrell JS (1999) Probability-based protein identification by searching sequence databases using mass spectrometry data. Electrophoresis 20:551–567
Lee CF, Griffiths S, Rodríguez-Suárez E, Pierce A, Unwin RD, Jaworska E, Evans CA, Gaskell SJ, Whetton AD (2010) Assessment of downstream effectors of BCR/ABL protein tyrosine kinase using combined proteomic approaches. Proteomics 10:3321–3342
Pandey A, Podtelejnikov AV, Blagoev B, Bustelo XR, Mann M, Lodish HF (2000) Analysis of receptor signaling pathways by mass spectrometry: identification of vav-2 as a substrate of the epidermal and platelet-derived growth factor receptors. Proc Natl Acad Sci U S A 97:179–184
Gronborg M, Kristiansen TZ, Stensballe A, Andersen JS, Ohara O, Mann M, Jensen ON, Pandey A (2002) A mass spectrometry-based proteomic approach for identification of serine/threonine-phosphorylated proteins by enrichment with phospho-specific antibodies: identification of a novel protein, Frigg, as a protein kinase A substrate. Mol Cell Proteomics 1:517–527
Kim SC, Sprung R, Chen Y, Xu Y, Ball H, Pei J, Cheng T, Kho Y, Xiao H, Xiao L, Grishin NV, White M, Yang XJ, Zhao Y (2006) Substrate and functional diversity of lysine acetylation revealed by a proteomics survey. Mol Cell 23:607–618
Choudhary C, Kumar C, Gnad F, Nielsen ML, Rehman M, Walther TC, Olsen JV, Mann M (2009) Lysine acetylation targets protein complexes and co-regulates major cellular functions. Science 325:834–840
Unwin R, Griffiths J, Leverentz M, Grallert A, Hagan I, Whetton A (2005) Multiple reaction monitoring to identify sites of protein phosphorylation with high sensitivity. Mol Cell Proteomics 4:1134–1144
Acknowledgments
Financial support from Leukaemia Lymphoma Research, UK (ADW, CE, JG), Biotechnology and Biological Sciences Research Council (BC), Cancer Research, UK (ADW, JRG), and NIHR Manchester Biomedical Research Centre (RDU). Financial support from Engineering and Physical Sciences Research Council, ChELSI initiative EP/E036252/1 (CE).
Author information
Authors and Affiliations
Editor information
Editors and Affiliations
Rights and permissions
Copyright information
© 2013 Springer Science+Business Media, LLC
About this protocol
Cite this protocol
Evans, C.A., Griffiths, J.R., Unwin, R.D., Whetton, A.D., Corfe, B.M. (2013). Application of the MIDAS Approach for Analysis of Lysine Acetylation Sites. In: Hake, S., Janzen, C. (eds) Protein Acetylation. Methods in Molecular Biology, vol 981. Humana Press, Totowa, NJ. https://doi.org/10.1007/978-1-62703-305-3_3
Download citation
DOI: https://doi.org/10.1007/978-1-62703-305-3_3
Published:
Publisher Name: Humana Press, Totowa, NJ
Print ISBN: 978-1-62703-304-6
Online ISBN: 978-1-62703-305-3
eBook Packages: Springer Protocols