Hello. I have been struggling with purification of human Tyrosinase expressed in E. Coli. In brief, I induce expression in E, Coli BL21 at an OD of 0.6 and then incubate further for 3 hours at 37 degrees or overnight at 18 degrees. In both cases I seem to have problems with expressing enough protein. I am attributing this to low cell density. I have read several literature and I have seen some labs use bioreactors which I don’t have. Can someone who has expressed human Tyrosinase in E. Coli BL21 (DE3) recommend an appropriate protocol that doesn’t involve use of bioreactors or fed-batch technology?
cell density only affects the total amount of protein you harvested, not the protein amount of each cell produced. In fact, since you induced at very low OD, each cell should produce a lot of your protein if the expression system works well. Did you see a strong band of your protein relative to other proteins in SDS gel?
If you worry total protein amount, you can let OD reaches 1.5 or even 1.8 and then start induction. In LB, you still have sufficient amount of nutrition left at this OD for synthesis of your target protein
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