'We describe a capillary electrophoresis (CE) assay to detect G protein-coupled receptor (GPCR)-stimulated G protein GTPase activity in cell membranes expressing alpha2A adrenoreceptor-Galphao1 wild-type (wt) or C351I mutant fusion proteins using a fluorescent, hydrolyzable GTP analogue. As no change in total fluorescence is observed by conversion of substrate to product, ... More
Automated single-cell electroporation.
AuthorsBae C, Butler PJ
JournalBiotechniques
PubMed ID17068953
Identification of a small GTPase inhibitor using a high-throughput flow cytometry bead-based multiplex assay.
AuthorsSurviladze Z, Waller A, Wu Y, Romero E, Edwards BS, Wandinger-Ness A, Sklar LA,
JournalJ Biomol Screen
PubMed ID20008126
Small GTPases are key regulators of cellular activity and represent novel targets for the treatment of human diseases using small-molecule inhibitors. The authors describe a multiplex, flow cytometry bead-based assay for the identification and characterization of inhibitors or activators of small GTPases. Six different glutathione-S-transferase (GST)-tagged small GTPases were bound ... More
A direct fluorescence-based assay for RGS domain GTPase accelerating activity.
AuthorsWillard FS, Kimple AJ, Johnston CA, Siderovski DP
JournalAnal Biochem
PubMed ID15840508
Diverse extracellular signals regulate seven transmembrane-spanning receptors to modulate cellular physiology. These receptors signal primarily through activation of heterotrimeric guanine nucleotide binding proteins (G proteins). A major determinant of heterotrimeric G protein signaling in vivo and in vitro is the intrinsic GTPase activity of the Galpha subunit. RGS (regulator of ... More
Real-time detection of basal and stimulated G protein GTPase activity using fluorescent GTP analogues.
Hydrolysis of fluorescent GTP analogues BODIPY FL guanosine 5 '-O-(thiotriphosphate) (BGTPgammaS) and BODIPY FL GTP (BGTP) by Galpha(i1) and Galpha was characterized using on-line capillary electrophoresis (o) laser-induced fluorescence assays in order that changes in sub-strate, substrate-enzyme complex, and product could be monitored separately. Apparent k values (V /[E]) (max ... More
Cholesterol sulfate is a DOCK2 inhibitor that mediates tissue-specific immune evasion in the eye.
Authors
JournalSci Signal
PubMed ID30065028
A kinetic safety gate controlling the delivery of unnatural amino acids to the ribosome.
Authors
JournalJ Am Chem Soc
PubMed ID24079513
Targeting Ras-Driven Cancer Cell Survival and Invasion through Selective Inhibition of DOCK1.