Skip to main content
SpringerLink
Log in

Phosphotyrosine Profiling Using SILAC

  • Protocol
  • First Online:
SILAC

Part of the book series: Methods in Molecular Biology ((MIMB,volume 2603))

  • 724 Accesses

Abstract

Tyrosine phosphorylation on proteins is an important posttranslational modification that regulates various processes in cells. Mass spectrometry-based phosphotyrosine profiling can reveal tyrosine kinase signaling activity in cells. Using quantitative proteomics strategies such as stable isotope labeling with amino acids in cell culture (SILAC) allows comparison of tyrosine kinase signaling activity across two to –three different conditions. In this book chapter, we discuss the reagents required and a step-by-step protocol to carry out phosphotyrosine profiling using SILAC.

This is a preview of subscription content, log in via an institution to check access.

Access this chapter

Log in via an institution
Protocol
USD 49.95
Price excludes VAT (USA)
  • Available as PDF
  • Read on any device
  • Instant download
  • Own it forever
eBook
USD 109.00
Price excludes VAT (USA)
  • Available as EPUB and PDF
  • Read on any device
  • Instant download
  • Own it forever
Softcover Book
USD 139.99
Price excludes VAT (USA)
  • Compact, lightweight edition
  • Dispatched in 3 to 5 business days
  • Free shipping worldwide - see info
Hardcover Book
USD 199.99
Price excludes VAT (USA)
  • Durable hardcover edition
  • Dispatched in 3 to 5 business days
  • Free shipping worldwide - see info

Tax calculation will be finalised at checkout

Purchases are for personal use only

Institutional subscriptions

References

  1. Hornbeck PV, Kornhauser JM, Tkachev S, Zhang B, Skrzypek E, Murray B, Latham V, Sullivan M (2012) PhosphoSitePlus: a comprehensive resource for investigating the structure and function of experimentally determined post-translational modifications in man and mouse. Nucleic Acids Res 40:D261–D270

    Article  CAS  PubMed  Google Scholar 

  2. Pottier C, Fresnais M, Gilon M, Jérusalem G, Longuespée R, Sounni NE (2020) Tyrosine kinase inhibitors in cancer: breakthrough and challenges of targeted therapy. Cancer 12:731

    Article  CAS  Google Scholar 

  3. Syed N, Barbhuiya MA, Pinto SM, Nirujogi RS, Renuse S, Datta KK, Khan AA, Srikumar K, Prasad TS, Kumar MV, Kumar RV, Chatterjee A, Pandey A, Gowda H (2015) Phosphotyrosine profiling identifies ephrin receptor A2 as a potential therapeutic target in esophageal squamous-cell carcinoma. Proteomics 15:374–382

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  4. Sathe G, Pinto SM, Syed N, Nanjappa V, Solanki HS, Renuse S, Chavan S, Khan AA, Patil AH, Nirujogi RS, Nair B, Mathur PP, Prasad TSK, Gowda H, Chatterjee A (2016) Phosphotyrosine profiling of curcumin-induced signaling. Clin Proteomics 13:13

    Article  PubMed  PubMed Central  Google Scholar 

  5. Kretschmar C (1990) Alzheimer’s disease. Course and therapeutic possibilities. Deutsche Krankenpflegezeitschrift 43:8–10

    CAS  PubMed  Google Scholar 

  6. van der Mijn JC, Labots M, Piersma SR, Pham TV, Knol JC, Broxterman HJ, Verheul HM, Jiménez CR (2015) Evaluation of different phospho-tyrosine antibodies for label-free phosphoproteomics. J Proteome 127:259–263

    Article  Google Scholar 

  7. Harsha HC, Molina H, Pandey A (2008) Quantitative proteomics using stable isotope labeling with amino acids in cell culture. Nat Protoc 3:505–516

    Article  CAS  PubMed  Google Scholar 

  8. Datta KK, Madugundu AK, Gowda H (2016) Proteogenomic methods to improve genome annotation. Methods Mol Biol 1410:77–89

    Article  CAS  PubMed  Google Scholar 

  9. Patil AH, Datta KK, Behera SK, Kasaragod S, Pinto SM, Koyangana SG, Mathur PP, Gowda H, Pandey A, Prasad TSK (2018) Dissecting candida pathobiology: post-translational modifications on the Candida tropicalis Proteome. OMICS J Integr Biol 22:544–552

    Article  CAS  Google Scholar 

Download references

Author information

Authors and Affiliations

  1. Department of Genetics and Computational Biology, QIMR Berghofer Medical Research Institute, Brisbane, QLD, Australia

    Keshava K. Datta & Harsha Gowda

  2. Institute of Bioinformatics, Bangalore, India

    Aditi Chatterjee

  3. School of Biomedical Sciences, The University of Queensland, Brisbane, QLD, Australia

    Harsha Gowda

  4. Faculty of Health, Queensland University of Technology, Brisbane, QLD, Australia

    Harsha Gowda

Authors
  1. Keshava K. Datta
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Aditi Chatterjee
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Harsha Gowda
    View author publications

    You can also search for this author in PubMed Google Scholar

Editor information

Editors and Affiliations

  1. Analytical Chemistry Department, Faculty of Chemical Sciences, Complutense University of Madrid, Madrid, Spain

    Jose L. Luque-Garcia

Rights and permissions

Reprints and permissions

Copyright information

© 2023 The Author(s), under exclusive license to Springer Science+Business Media, LLC, part of Springer Nature

About this protocol

Check for updates. Verify currency and authenticity via CrossMark

Cite this protocol

Datta, K.K., Chatterjee, A., Gowda, H. (2023). Phosphotyrosine Profiling Using SILAC. In: Luque-Garcia, J.L. (eds) SILAC. Methods in Molecular Biology, vol 2603. Humana, New York, NY. https://doi.org/10.1007/978-1-0716-2863-8_9

Download citation

  • DOI: https://doi.org/10.1007/978-1-0716-2863-8_9

  • Published:

  • Publisher Name: Humana, New York, NY

  • Print ISBN: 978-1-0716-2862-1

  • Online ISBN: 978-1-0716-2863-8

  • eBook Packages: Springer Protocols

Publish with us

Policies and ethics

Search

Navigation