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| | PqsA Spectrophotometric Assay |
| Description: | Reactions were performed at 37 °C in a 0.5 mL volume in a Varian Cary 100 UV-visible spectrophotometer with Cary WinUV software. Reaction mixtures contained 100 mM HEPES, pH 8.0, 0.2 mM dithiothreitol, 2 mM MgCl2, ~3.3 μg PqsA protein (~60 nM final), and depending on which substrate varied, 1 mM ATP (disodium salt), 0.5 mM coenzyme A (trilithium salt), 0.5 mM sodium anthranilate, and inhibitor (usually as a DMSO stock solution). Reactions were blanked and preincubated with all components except anthranilate at 37 °C for 1 min, then initiated by addition of anthranilate. Formation of anthraniloyl-CoA was monitoredby absorbance at 365 nm (ε = 5.5 mM-1cm-1) for 1 min. |
| Affinity data for this assay | |
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