GST pull down assay

Mix the GST tagged protein and fluorescent protein with 30 μl Glutathione Sepharose beads (Cytiva) at final 1 μM concentration in the buffer of 25 mM HEPES at pH 7.5, 150 mM NaCl, 1 mM MgCl2 and 1 mM TCEP. The total volume is 200 μl.

Rocking at 4°C

Wash the beads four times, then eluted in 50 μl of buffer with 25 mM glutathione

Mix 18 μl eluent in lithium dodecylsulfate (LDS)/BME buffer and subjected to SDS/PAGE gel without heating samples. The gel was scanned at 488 or 550 nm in ChemiDoc MP imaging system (Bio-Rad).