Astra 6.1 program

We carried out the SEC shift assay to analyze the protein–DNA interactions. The BAZ2A (2 mg/ml) protein was mixed with dsDNA in a molar ratio of 1:1.1 and incubated on ice for 30 min. The samples and the protein molecular weight standard sample were analyzed by size-exclusion chromatograms (Superdex 75 10/300GL, GE) using the buffer containing 20 mM Tris HCl, pH 7.5, 1 mM DTT, and 150 mM or 450 mM NaCl. The SEC elution fractions corresponding to each peak were analyzed by SDS-PAGE.
For molar weight determination of the protein–DNA complex, the SEC-MALS experiment was performed using an HPLC-MALS system. A DAWN TREOS multiangle light scattering detector (Wyatt Technology) and an Optilab T-rEX refractometer (Wyatt Technology) were used in-line with WTC-015S5 hydrophilic film bonded silica column (Wyatt Technology) pre-equilibrated in a PBS buffer containing 50 mM Na₂HPO₄, 50 mM NaH₂PO₄, and 50 mM NaCl, pH 6.8, at a flow rate of 0.5 ml/min. Molecular weights of proteins were calculated with a dn/dc value (refractive index increment) of 0.185 ml/g using the Astra 6.1 program (Wyatt Technology).

For molar mass determination, purified proteins or protein complexes were analyzed using an ÄKTA-MALS system. Note that 200 μg of protein solution was injected on Superdex 200 10/300 GL (GE Healthcare) equilibrated in 10 mM Tris 7.5, 0.5 mM DTT, 0.5 mM EDTA (pH 8.0), and 2 M NaCl at a flow rate of 0.3 mL per minute. Separation and ultraviolet detection were performed using an ÄKTA system (GE Healthcare), light scattering was monitored using a miniDAWN TREOS system (Wyatt Technology), and concentration was measured by an Optilab T-rEX differential refractometer (Wyatt Technology). Molar masses of proteins were calculated using the Astra 6.1 program (Wyatt Technology) with a dn/dc value of 0.185 mL/g.