Protein expression and purification

To generate HisMBP-SWI1^1-300, HisMBP-SWI1^301-639, HisGST-PDS5A^1-809 and HisMBP-WAPL1 constructs, the respective coding sequences were amplified by PCR and subcloned into pDONR223 vector by gateway BP reactions (Table ^S2). The resulting constructs were integrated by gateway LR reactions into pHMGWA or pHGGWA vectors for the His MBP- or HisGST- tagged fusions, respectively. For the heterologous expression, the constructs were transformed into the E. coli BL21 (DE3)pLysS cells and grown at 37 °C in the presence of 100 mg/l ampicillin until the OD₆₀₀ of 0.6. Protein expression was induced by adding IPTG to a final concentration of 0.3 mM, and the cells were incubated at 37 °C for 3 h (HisMBP-SWI1^301-639) or 18 °C overnight (HisMBP-SWI1^1-300, HisGST-PDS5A^1-809 and HisMBP-WAPL1). All proteins were purified under native conditions by using Ni-NTA sepharose (QIAGEN) according to the manual.

For the purification of PDS5A^1-809-WAPL1 heterodimers, the ampicillin resistance gene of WAPL1-pHMGWA was first replaced by the kanamycin resistance gene and co-transformed into BL21 (DE3)pLysS cells containing the vector PDS5A^1-809-pHGGWA. The cells harboring both constructs were grown at 37 °C in the presence of 100 mg/l ampicillin and 50 mg/l kanamycin until the OD₆₀₀ to 0.6 and induced with 0.3 mM IPTG at 18 °C for overnight. Cells were harvested and PDS5A^1-809 and WAPL1 heterodimers were purified using GST agarose beads (Novogene). Coomassie Brilliant Blue (CBB) stained gels of all purified proteins used in this study are shown in Supplementary Fig. ¹⁵. The protease inhibitor cocktail (Roche) was always used in the purification procedures.