The specific binding of small molecule isoprenoids to rhoGDP dissociation inhibitor (rhoGDI).
The activities of small G-proteins are in part regulated by their interactions with GDI proteins. This binding is thought to be dependent on the C-terminal isoprenoid modification (geranylgeranyl or farnesyl) of these proteins. G-proteins are generally isoprenylated/methylated at their C-terminal cysteine residues. A quantitative fluorescence assay is reported here to evaluate the specificity of binding of rhoGDI. A rhodamine-labeled geranylgeranylated/methylated cysteine derivative is used to measure its binding to rhoGDI. Saturable binding in the low micromolar range is found with various geranylgeranylated/farnesylated analogues. Interestingly, the carboxymethylated derivatives bound significantly better than their free acid counterparts, suggesting that the state of methylation of the analogues is important for binding. The binding is also selective with respect to isoprenoid. Analogues containing hydrophobic modifications other than geranylgeranyl or farnesyl do not bind with significant affinities. These data demonstrate a substantial degree of specificity in the binding of isoprenoids to a protein important in signal transduction.
Mondal
M S
MS
Department of Biological Chemistry and Molecular Pharmacology, Harvard Medical School, 240 Longwood Avenue, Boston, Massachusetts 02115, USA.
Wang
Z
Z
Seeds
A M
AM
Rando
R R
RR
D016880:Anisotropy; D003545:Cysteine / Q000031:analogs & derivatives* / Q000737:chemistry; D004224:Diterpenes / Q000737:chemistry*; D020730:Guanine Nucleotide Dissociation Inhibitors / Q000737:chemistry*; D008745:Methylation; D015394:Molecular Structure; D011485:Protein Binding; D015398:Signal Transduction; D020741:rho GTP-Binding Proteins / Q000737:chemistry; D062947:rho-Specific Guanine Nucleotide Dissociation Inhibitors
D004224:Diterpenes; D020730:Guanine Nucleotide Dissociation Inhibitors; D062947:rho-Specific Guanine Nucleotide Dissociation Inhibitors; C065109:geranylgeranylcysteine methyl ester; D020741:rho GTP-Binding Proteins; D003545:Cysteine