Culture pellets were resuspended in 50 mM Tris pH: 7.5, 300 mM NaCl and 5 µM β-mercaptoethanol (BME) with the addition of complete protease inhibitor (1 tab per 50 mL of lysis buffer). Following resuspension, lysis was performed on ice by sonication for 10 min (30 s on/ 30 s off, 45° amplitude). Clarification was then performed by centrifugation 1 hr at 12,000 g/4°C after which the supernatant was supplemented with 20 mM imidazole and further incubated with 5 ml Ni-NTA resin with a stirring magnet at 4°C for 30 min. Resin retention was performed by gravity with a Bio-Rad glass column after which the resin was washed with 100 mL of washing buffer (50 mM Tris pH: 7.5, 1M NaCl, 2 mM BME and 20 mM Imidazole). His-tagged TgCPSF4-YTH was then eluted by in 50 mM Tris pH: 7.5, 300 mM NaCl, 300 mM Imidazole, 2% glycerol and 2 mM BME and dialyzed overnight with TEV protease to remove the 8*histidine tag in a buffer composed of 50 mM Tris pH 7.5, 150 mM NaCl, 2% glycerol and 2 mM BME. Non-cleaved forms of TgCPSF4-YTH were removed by flowing through 1 mL of pre-equilibrated Ni-NTA. All subsequent liquid chromatography steps were performed on an Akta Purifier. Nucleic acid contaminants were removed by directly binding CPSF4-YTH onto a 5 ml heparin (GE healthcare) and eluting by a 40 ml step NaCl gradient (150 mM to 2M) in a 50 mM tris pH 7.5, 2 mM BME, 2% glycerol buffer system. Following elution from heparin, fractions of interest were pooled and concentrated to 1 mL using 10 kDa cut-off concentrators before being subsequently injected on an S75 column for size fractionation in a buffer containing 50 mM Tris pH: 7.5, 150 mM NaCl, 1 mM BME. Following the size exclusion step, final fractions were pooled, concentrated to a minimum of 15 mg/mL with centricon 10 kDa concentrators, flash frozen in liquid nitrogen and stored at −80°C.

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