In vitro translation system with PTM's

[Dkav]

I am having major issues with cell culture and i came up with this idea. Is it feasible to purify the necessary components of post translation modifications (golgi, er, and enzymes) and throw them into an in-vitro cell free translation system to make fully functional post translationally modified eukaryotic proteins? Thanks.

[relaxin]

I don't think so, some enzymes in the ER lumen are membrane bound and proprotein convertases are present only in secretary vessicles. It will be difficult to mimic normal cellular posttranslational modification in a cell-free translation system.

[Dkav]

I think some of the enzymes will still be attached to the membrane after gentle purification, no? ...and can't i just put proprotein convertase containing vesicles in my reaction mix?

[CrowSan]

Something similar to this already exists. I used Rabbit reticular lysate a few decades ago to made protein "in vitro". I imagine someone still sells this product. I'm not sure whether this system also does the full scale post-translational modifications or not though....

[mehedi107]

It will be difficult to mimic normal cellular posttranslational modification in a cell-free translation system.