Optimization of metabolic labeling with Alkyne-containing isoprenoid probes

Mina Ahmadi, Kiall Francis Suazo, Mark D. Distefano

Research output: Chapter in Book/Report/Conference proceedingChapter

2 Scopus citations

Abstract

Protein prenylation, found in eukaryotes, is a posttranslational modification in which one or two isoprenoid groups are added to the C terminus of selected proteins using either a farnesyl group or a geranylgeranyl group. Prenylation facilitates protein localization mainly to the plasma membrane where the prenylated proteins, including small GTPases, mediate signal transduction pathways. Changes in the level of prenylated proteins may serve a critical function in a variety of diseases. Metabolic labeling using modified isoprenoid probes followed by enrichment and proteomic analysis allows the identities and levels of prenylated proteins to be investigated. In this protocol, we illustrate how the conditions for metabolic labeling are optimized to maximize probe incorporation in HeLa cells through a combination of in-gel fluorescence and densitometric analysis.

Original languageEnglish (US)
Title of host publicationMethods in Molecular Biology
PublisherHumana Press Inc.
Pages35-43
Number of pages9
DOIs
StatePublished - 2019

Publication series

NameMethods in Molecular Biology
Volume2009
ISSN (Print)1064-3745
ISSN (Electronic)1940-6029

Bibliographical note

Funding Information:
This work was supported in part by the National Institutes of Health (RF1AG056976 and GM084152) and by the National Science Foundation grant (CHE-1308655).

Publisher Copyright:
© Springer Science+Business Media, LLC, part of Springer Nature 2019.

Keywords

  • Farnesylation
  • Geranylgeranylation
  • In-gel fluorescence
  • Isoprenoid analog
  • Lovastatin
  • Metabolic labeling
  • Protein prenylation

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