Mapping of Ras-related GTP-binding proteins by GTP overlay following two-dimensional gel electrophoresis.
Open accessAbstract
For identification of Rab, Rac, Rho, Ral, Rap, and Arf proteins on two-dimensional polyacrylamide gels, we have expressed full-length cDNAs of members of these protein families with the T7 RNA polymerase-recombinant vaccinia virus expression system. Membrane preparations from cells expressing the cDNAs were subjected to high-resolution two-dimensional polyacrylamide gel electrophoresis followed by [alpha-32P]GTP ligand blotting. We have mapped 28 small GTP-binding proteins relative to their isoelectric points and according to their molecular weights and by immunoblotting with specific antibodies. Rab and Rho proteins could be specifically identified by extraction of streptolysin O-permeabilized Madin-Darby canine kidney (MDCK) cells with Rab- and Rho-GDP dissociation inhibitor. We applied the reference mapping to analyze the GTP-binding patterns of synaptosome fractions from rat brain. The purified synaptosomes exhibited specific enrichment of Rab3a, Rab5a, Ral, and several other GTPases. This approach and the map we have produced should provide a useful aid for the analysis of the expression and localization of members of all families of small GTP-binding proteins in various cell types and subcellular fractions.
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Ral is expressed in the nervous system.
“Like the exocyst, Ral is expressed in the nervous system (Ngsee et al, 1991;Huber et al, 1994;Peng et al, 2004;Han et al, 2009).”
From Ral mediates activity‐dependent growth of postsynaptic membranes via recruitment of the exocyst (2013) · cites this paper as Huber et al, 1994
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- [46]Chen, Y.-T., Holcomb, C. & Moore, H.-P. H. (1993) Proc. Natl. Acad. Sci. USA 90, 6508-6512.
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Seabra, M. C., Ho, Y. K., & Anant, J. S. (1995) · Journal of Biological Chemistry · doi:10.1074/jbc.270.41.24420 · Cited by 223 · see citing works
Ral mediates activity‐dependent growth of postsynaptic membranes via recruitment of the exocyst
Teodoro, R. O., Pekkurnaz, G., Nasser, A., Higashi‐Kovtun, M. E., Balakireva, M., McLachlan, I. G., Camonis, J., & Schwarz, T. L. (2013) · The EMBO Journal · doi:10.1038/emboj.2013.147 · Cited by 61 · see citing works
- Authors
- Huber, L. A., Ullrich, O., Takai, Y., Lütcke, A., Dupree, P., Olkkonen, V, Virta, H., de Hoop, M. J., Alexandrov, K., & Peter, M.
- Affiliations
- European Molecular Biology Laboratory (DE)
- European Molecular Biology Laboratory, Cell Biology Programme, Heidelberg, Germany.
- Venue
- Proceedings of the National Academy of Sciences
- Publisher
- National Academy of Sciences
- Published
- 1994-08-16
- Volume
- 91
- Issue
- 17
- Pages
- 7874-7878
- ISSN
- 0027-8424 (ISSN-L)
- 1091-6490
- PMC
- PMC44506
- OpenAlex
- W2170786874
- Cited by
- 50
- References
- 0
- Citations by year
- 2023: 1 · 2013: 3 · 2012: 1
- FWCI
- 1.90
- Type
- journal article
- Language
- en
- Open access
- green
- europepmc.org
- Locations
- Proceedings of the National Academy of Sciences (publishedVersion)
- Europe PMC (PubMed Central) (publishedVersion)
- PubMed Central (publishedVersion)
- PubMed (publishedVersion)
- QUT ePrints (Queensland University of Technology) (submittedVersion)
- Funders
- Deutsche Forschungsgemeinschaft
- Topics
- Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › Protein Kinase Regulation and GTPase Signaling
- Life Sciences › Biochemistry, Genetics and Molecular Biology › Cell Biology › Cellular transport and secretion
- Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › Fungal and yeast genetics research
- Keywords
- small GTPases, Rab GTPases, Rho proteins, Arf proteins, protein mapping, synaptosomes, Rab3A, Rab5a, Rho GDP dissociation inhibitor, vaccinia virus expression system, subcellular localization
- MeSH terms
- Animals, Autoradiography, Cell Line, Cell Membrane, Cells, Cultured, Dogs, Electrophoresis, Polyacrylamide Gel, Guanosine Triphosphate, Cricetinae, Hippocampus, Kidney, Molecular Weight, Neurons, Synaptosomes, Immunoblotting, Electrophoresis, Gel, Two-Dimensional, GTP-Binding Proteins, Rats
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