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The temperature was kept at 300 K in all simulations using a velocity rescaling thermostat 36 , and the pressure was kept at 1 bar (using a Berendsen barostat 37 for the equilibration steps and a ParrinelloRahman barostat 38 for the production)

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Central to this advancement is the innovative use of the Tiron/ferrous ion (Fe 2+ ) reagent, which simplifies the assay by eliminating the need for protein precipitation, a step that previously introduced variability