ArticleCombinatorial chemistry & high throughput screening2008
A high throughput fluorescence polarization assay for inhibitors of the GoLoco motif/G-alpha interaction.
Article in Combinatorial chemistry & high throughput screening, 2008. The graph could read no effect estimate from its abstract, so it casts no vote on the map. Cited by 21 papers.
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Who cites it
21 citing papers in PubMed, 29 citations in OpenAlex.
- Article
- Astrocytes as Guardians of Neuronal Excitability: Mechanisms Underlying Epileptogenesis.Frontiers in neurology · 2020Review
- Regulator of G-protein signaling (RGS) proteins as drug targets: Progress and future potentials.The Journal of biological chemistry · 2019Review
- Specific inhibition of GPCR-independent G protein signaling by a rationally engineered protein.Proceedings of the National Academy of Sciences of the United States of America · 2017Article
- Fluorescence polarization assays to measure interactions between Gα subunits of heterotrimeric G proteins and regulatory motifs.Methods in cell biology · 2017Article
- Fluorescence polarization assays in high-throughput screening and drug discovery: a review.Methods and applications in fluorescence · 2016Article
- Heterotrimeric G protein signaling via GIV/Girdin: Breaking the rules of engagement, space, and time.BioEssays : news and reviews in molecular, cellular and developmental biology · 2016Review
- GIV/Girdin transmits signals from multiple receptors by triggering trimeric G protein activation.The Journal of biological chemistry · 2015Review
- Article
- Cullin3-BTB interface: a novel target for stapled peptides.PloS one · 2015Article
- Regulator of G-protein signaling-21 (RGS21) is an inhibitor of bitter gustatory signaling found in lingual and airway epithelia.The Journal of biological chemistry · 2012Article
- Identification of influenza endonuclease inhibitors using a novel fluorescence polarization assay.ACS chemical biology · 2012Article
- A P-loop mutation in Gα subunits prevents transition to the active state: implications for G-protein signaling in fungal pathogenesis.PLoS pathogens · 2012Article
- Regulators of G-protein signaling and their Gα substrates: promises and challenges in their use as drug discovery targets.Pharmacological reviews · 2011Review
- G Protein binding sites on Calnuc (nucleobindin 1) and NUCB2 (nucleobindin 2) define a new class of G(alpha)i-regulatory motifs.The Journal of biological chemistry · 2011Article
- Structural determinants of affinity enhancement between GoLoco motifs and G-protein alpha subunit mutants.The Journal of biological chemistry · 2011Article
- Fluorescence polarization assays in small molecule screening.Expert opinion on drug discovery · 2011Article
- A homogeneous method to measure nucleotide exchange by α-subunits of heterotrimeric G-proteins using fluorescence polarization.Assay and drug development technologies · 2010Article
- A capture coupling method for the covalent immobilization of hexahistidine tagged proteins for surface plasmon resonance.Methods in molecular biology (Clifton, N.J.) · 2010Article
- Ligand-based peptide design and combinatorial peptide libraries to target G protein-coupled receptors.Current pharmaceutical design · 2010Review
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Authors and funding
11 authors at 3 institutions in 2 countries.
Funding
Abstract
The GoLoco motif is a short Galpha-binding polypeptide sequence. It is often found in proteins that regulate cell-surface receptor signaling, such as RGS12, as well as in proteins that regulate mitotic spindle orientation and force generation during cell division, such as GPSM2/LGN. Here, we describe a high throughput fluorescence polarization (FP) assay using fluorophore-labeled GoLoco motif peptides for identifying inhibitors of the GoLoco motif interaction with the G-protein alpha subunit Galpha (i1). The assay exhibits considerable stability over time and is tolerant to DMSO up to 5%. The Z'-factors for robustness of the GPSM2 and RGS12 GoLoco motif assays in a 96-well plate format were determined to be 0.81 and 0.84, respectively; the latter assay was run in a 384-well plate format and produced a Z'-factor of 0.80. To determine the screening factor window (Z-factor) of the RGS12 GoLoco motif screen using a small molecule library, the NCI Diversity Set was screened. The Z-factor was determined to be 0.66, suggesting that this FP assay would perform well when developed for 1,536-well format and scaled up to larger libraries. We then miniaturized to a 4 microL final volume a pair of FP assays utilizing fluorescein- (green) and rhodamine- (red) labeled RGS12 GoLoco motif peptides. In a fully-automated run, the Sigma-Aldrich LOPAC(1280) collection was screened three times with every library compound being tested over a range of concentrations following the quantitative high throughput screening (qHTS) paradigm; excellent assay performance was noted with average Z-factors of 0.84 and 0.66 for the green- and red-label assays, respectively.
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Registered trials
Read under generation 80e0d062 · epoch 390. Bibliography from PubMed, PubMed Central and OpenAlex; grants from NIH RePORTER; trial links from ClinicalTrials.gov; estimates, votes and beliefs from the OpenQuestion graph.