Expression of TFAAA-USP18 in pSUMO vector backbone under different conditions(A) TFAAA-USP18 was expressed in E. coli BL21(DE3)pLysS at 37°C. Expression was verified by analyzing protein content directly after lysis on SDS-PAGE followed by Coomassie staining. After 3 h of induction TFAAA-USP18 fusion protein made up more than 50% of whole cellular proteins. (B) Soluble and insoluble fractions from (A) were analysed by Western blot with an anti His6-Tag antibody. Almost all fusion protein was present in the insoluble fraction and only a faint band for soluble protein was observed. (C) Expression of TFAAA-USP18 at 15°C in E. coli BL21(DE3)pLysS yielded soluble protein. Western blot analysis using an anti His6-Tag antibody detected TFAAA-USP18 only in the soluble fraction. (D) E. coli Tuner(DE3) and E. coli Tuner(DE3)pLysS were tested for expression of TFAAA-USP18 at 15°C. Soluble and insoluble fractions were analysed by SDS-PAGE and subsequent Coomassie staining. Strong expression was only observed in E. coli Tuner(DE3). The major portion of the fusion protein was observed in the soluble fraction.
Basters et al. BMC Biotechnology 2012 12:56 doi:10.1186/1472-6750-12-56