Abstract
S-acylation is increasingly being recognized as an important posttranslational modification of proteins controlling activity, subcellular localization, microdomain residence, and stability. Heterotrimeric G-proteins and GPCRs are particularly well studied S-acylated proteins, and fast, cheap, reliable methods are required for the analysis of S-acylation states of these proteins. Various approaches have been developed to study S-acylation, but they are time consuming, expensive, frequently require radiolabels and generally only suitable for cell culture, making them impractical for work in plant systems. Here a rapid and inexpensive method is described for the analysis of the S-acylation state of AGG2 that can be performed on any cell or tissue sample using standard laboratory equipment and methods. This method is also applicable to any protein that can be detected by western blotting.
Access this chapter
Tax calculation will be finalised at checkout
Purchases are for personal use only
References
Ovchinnikov YA, Abdulaev NG, Bogachuk AS (1988) Two adjacent cysteine residues in the C-terminal cytoplasmic fragment of bovine rhodopsin are palmitylated. FEBS Lett 230:1–5
Adjobo-Hermans MJ, Goedhart J, Gadella TW Jr (2006) Plant G protein heterotrimers require dual lipidation motifs of Galpha and Ggamma and do not dissociate upon activation. J Cell Sci 119:5087–5097
Zeng Q, Wang X, Running MP (2007) Dual lipid modification of Arabidopsis Ggamma-subunits is required for efficient plasma membrane targeting. Plant Physiol 143:1119–1131
Running MP, Lavy M, Sternberg H, Galichet A, Gruissem W, Hake S, Ori N, Yalovsky S (2004) Enlarged meristems and delayed growth in plp mutants result from lack of CaaX prenyltransferases. Proc Natl Acad Sci USA 101:7815–7820
Hemsley PA, Taylor L, Grierson CS (2008) Assaying protein palmitoylation in plants. Plant Methods 4:2
Drisdel RC, Green WN (2004) Labeling and quantifying sites of protein palmitoylation. Biotechniques 36:276–285
Hou H, Subramanian K, LaGrassa TJ, Markgraf D, Dietrich LE, Urban J, Decker N, Ungermann C (2005) The DHHC protein Pfa3 affects vacuole-associated palmitoylation of the fusion factor Vac8. Proc Natl Acad Sci USA 102:17366–17371
Hemsley PA, Weimar T, Lilley KS, Dupree P, Grierson CS (2013) A proteomic approach identifies many novel palmitoylated proteins in Arabidopsis. New Phytol 197:805–814
Wessel D, Flugge UI (1984) A method for the quantitative recovery of protein in dilute solution in the presence of detergents and lipids. Anal Biochem 138:141–143
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
Hemsley, P.A. (2013). Assaying Protein S-Acylation in Plants. In: Running, M. (eds) G Protein-Coupled Receptor Signaling in Plants. Methods in Molecular Biology, vol 1043. Humana Press, Totowa, NJ. https://doi.org/10.1007/978-1-62703-532-3_15
Download citation
DOI: https://doi.org/10.1007/978-1-62703-532-3_15
Published:
Publisher Name: Humana Press, Totowa, NJ
Print ISBN: 978-1-62703-531-6
Online ISBN: 978-1-62703-532-3
eBook Packages: Springer Protocols