Monitoring wnt protein acylation using an in vitro cyclo- addition reaction

Rubina Tuladhar, Nageswari Yarravarapu, Lawrence Lum

Research output: Chapter in Book/Report/Conference proceedingChapter

Abstract

We describe here a technique for visualizing the lipidation status of Wnt proteins using azide-alkyne cycloaddition chemistry (click chemistry) and SDS-PAGE. This protocol incorporates in vivo labeling of a Wnt- IgG Fc fusion protein using an alkynylated palmitate probe but departs from a traditional approach by incorporating a secondary cycloaddition reaction performed on single-step purified Wnt protein immobilized on protein A resin. This approach mitigates experimental noise by decreasing the contribution of labeling from other palmitoylated proteins and by providing a robust method for normalizing labeling efficiency based on protein abundance.

Original languageEnglish (US)
Title of host publicationMethods in Molecular Biology
PublisherHumana Press Inc.
Pages11-16
Number of pages6
DOIs
StatePublished - Jan 1 2016

Publication series

NameMethods in Molecular Biology
Volume1481
ISSN (Print)1064-3745

Keywords

  • Biotinylation
  • Click chemistry
  • IgG fusion protein
  • Palmitoleate
  • Wnt acylation

ASJC Scopus subject areas

  • Molecular Biology
  • Genetics

Fingerprint Dive into the research topics of 'Monitoring wnt protein acylation using an in vitro cyclo- addition reaction'. Together they form a unique fingerprint.

  • Cite this

    Tuladhar, R., Yarravarapu, N., & Lum, L. (2016). Monitoring wnt protein acylation using an in vitro cyclo- addition reaction. In Methods in Molecular Biology (pp. 11-16). (Methods in Molecular Biology; Vol. 1481). Humana Press Inc.. https://doi.org/10.1007/978-1-4939-6393-5_2