Figure 6.

Native PAGE and qualitative SEC analyses to compare the stability of the in vivo-associated and renatured complexes of Scc4:His₆-Scc1. (A) Native PAGE of the in vivo-associated complex (lanes 1 and 2) and the renatured complex (lanes 3 and 4) after purification and after 2 days at room temperature. Lane 5 is Scc4 for comparison. Lane 6 is the Bio-Rad Precision Plus protein ladder with molecular weights listed to the right in kDa; the ladder was included as a qualitative guide. The full gel from which these lanes were extracted is shown in Figure S8. (B,C) Qualitative SEC analysis to assess the stability of (B) the in vivo-associated complex after purification (0 d, black solid line) and after 2 days at room temperature (dashed black line) and (C) the renatured complex after purification (0 d, gray solid line) and after 2 days at room temperature (dashed gray line). (C) The right vertical axis corresponds to the renatured complex after 2 days; a smaller amount was analyzed to reverse the bulk of the sample for NMR analysis. Differences in the elution times between the (B) in vivo- and (C) on-column-associated complexes are not interpretable (refer to Section 2.5 for details).