Gel Filtration Chromatography

Gel filtration chromatography was performed on BioLogic DuoFlow system (Bio-Rad) with a Superdex 200 column (GE Healthcare Life Sciences, Pittsburg, PA). 0.5 ml of purified Wzm proteins (250 μg) were injected into the column, which was equilibrated with the running buffer (150 mM NaCl, 50 mM sodium phosphate, 20% glycerol, 0.01% DDM, pH 8.0), and eluted at 0.5 ml per fraction (Fig. S1). Protein molecular weight standards, including Thyroglobulin (669 kDa, bovine), γ-Globulin (158 kDa, bovine), Ovalbumin (44 kDa, chicken) (Bio-Rad), and Apoferritin (443 kDa, horse spleen) (Sigma-Aldrich, St. Louis, MO) were separated by BioLogic DuoFlow system under the same conditions and confirmed by the UV detector of the system. The molecular weights of the Wzm protein complexes were estimated based on the linear regression of the molecular weight standards and their retention volumes.