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Ufc1 stimulates the rate of Uba5∼Ufm1 thioester formation. A, Uba5∼Ufm1 thioester levels were monitored over time using a Western blot-based assay. Uba5∼Ufm1 formation was quantified in reaction mixtures containing buffer (●), wild-type Ufc1 (■), C116A Ufc1(▴), or purified Ufc1∼Ufm1(♦). B, rates of Uba5∼Ufm1 formation were determined by linear regression during the linear range and plotted to scale.
