| Assay Method Information | |
| | Isothermal Titration Calorimetry (ITC) |
| Description: | ITC titrations were performed using an ITC200 instrument (Microcal Inc.,Malvern). DMSO stock solutions (20 mM) of fragments were diluted 1:20 (v/v) into ITC titration buffer (50 mM tricine, 2 mM MnCl2, pH = 8.5) to give a 1 mM final compound concentration at 5% (v/v) DMSO. Aliquots of the same buffer batch (stored at −253 K) were used as for protein storage to avoid a buffer mismatch. H6PqsE was diluted in the same buffer to 100 μM, and DMSO concentration was adjusted to 5% (v/v). Titrations were carried out at 298 K using 19 injections of 2 μL every 180 s. Areas under the peaks were integrated. Baseline peaks at the endof a titration accounting for the heat of dilution and mixing were subtracted from the measurement. Data were fitted to a 1:1 binding model (MicroCal Origin 7 software). |
| Affinity data for this assay | |
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