Figure 6

SDS solubilization and purification of recombinant DGTA1 from insoluble fraction by Ni-NTA affinity chromatography. Recombinant DGAT1 was induced by IPTG for various time. Cell extract was centrifuged at 10,000g resulting in the supernatant (S10) and the pellet (P10). S10 was further centrifuged at 25,000g resulted in the supernatant (S25) and the pellet (P25). P10 and P25 were solubilized by 0.4% SDS followed by centrifugation at 50,000g (S50). S50 and Ni-NTA purification fractions were used for detecting rDGAT1 by immunoblotting with anti-MBP-hTTP antibodies. (A) Purification of rDGAT1 from SDS-solubilized fraction. S, the supernatant S25, U, unbound, W, wash, E, elution. (B) Time course of rDGAT1 purified from SDS-solubilized fraction. The full-length rDGAT1 is marked with an arrow.