Preparation of FRET reporters to support chemical probe development.
In high throughput screening (HTS) campaigns, the quality and cost of commercial reagents suitable for pilot studies often create obstacles upon scale-up to a full screen. We faced such challenges in our efforts to implement an HTS for inhibitors of the phosphopantetheinyl transferase Sfp using an assay that had been validated using commercially available reagents. Here we demonstrate a facile route to the synthetic preparation of reactive tetraethylrhodamine and quencher probes, and their application to economically produce fluorescent and quencher-modified substrates. These probes were prepared on a scale that would allow a full, quantitative HTS of more than 350,000 compounds.
Foley
Timothy L
TL
Department of Chemistry and Biochemistry, University of California-San Diego, 9500 Gilman Drive, La Jolla, CA 92093-0358, USA.
Yasgar
Adam
A
Garcia
Christopher J
CJ
Jadhav
Ajit
A
Simeonov
Anton
A
Burkart
Michael D
MD
D001426:Bacterial Proteins / Q000037:antagonists & inhibitors; D031541:Fluorescence Resonance Energy Transfer; D005456:Fluorescent Dyes / Q000138:chemical synthesis* / Q000737:chemistry; D047428:Protein Kinase Inhibitors / Q000737:chemistry; D012235:Rhodamines / Q000138:chemical synthesis* / Q000737:chemistry; D017855:Transferases (Other Substituted Phosphate Groups) / Q000037:antagonists & inhibitors
D001426:Bacterial Proteins; D005456:Fluorescent Dyes; D047428:Protein Kinase Inhibitors; D012235:Rhodamines; C082662:phosphopantetheinyl transferase; D017855:Transferases (Other Substituted Phosphate Groups); C029773:rhodamine B