Figure 3.
ssDNA promotes disulfide between catalytic nucleophiles. (A) Gel demonstrating the internal disulfide bond between C1 and C +1 and influence of ssDNA. Pho RadA C+1 was purified under nonreducing conditions (–TCEP). Reaction mixtures were incubated at the indicated temperatures for 5 min, separated by SDS–PAGE, and stained with Coomassie. Levels of the oxidized precursor (POX) and reduced precursor (PRED) were measured by densitometry. Tris-EDTA (TE) was used as a buffer control in the absence of ssDNA. (B) Quantification of C1–C+1 disulfide bond formation with or without ssDNA and TCEP prior to incubation (T0) or after 5 min at 63 °C. Error bars represent the standard deviation from three independent experiments.