Enzyme ATE1 plays role in cellularstressresponse, opening door to new therapeutic targets umbc NatureComms
. Briefly, the protein was brought into the anoxic chamber and allowed to equilibrate with the anaerobic chamber’s atmosphere overnight at 6 °C with shaking. Protein was then diluted to 100 μmol/L in reconstitution buffer comprising 25 mmol/L 3-morpholinopropane-1-sulfonic acid or 25 mmol/L Tris, pH 7.5, 300 mmol/L KCl, 1 mmol/L DTT, 0.7 mol/L glycerol. About 10 mmol/L stock FeClwas first titrated into the apoprotein until 4 mole eq. to 10 mole eq.
Circular dichroism spectra were recorded on a nitrogen-flushed spectropolarimeter operating at room temperature. Samples were contained within a quartz cuvette with a path length of 0.3 cm, and data were acquired from 800 to 300 nm with the instrument set to a spectral bandwidth of 1 nm. The data presented are an average of five scans.Samples containing ≈0.5 to 2 mmol/L iron in buffer plus 3.
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