Hi,
does anyone know of a database or method for predicting the expression of a recombinant protein in E. Coli which strain of E. coli is better to have the highest soluble expression of the protein predicted by the database, and which back-bone vector would be best for expression?
Thank you in advance for your help.
Hello! I saw MobiDB for Uniprot, but there are some very useful tools here, https://www.expasy.org/ You will most likely need to do expression testing. There are a few papers about this. BL21 is a decent standard E. coli strain for expression as is pET17b as a standard plasmid. There are a myriad of different expression plasmids and a few strains of E. coli. Induction conditions and growth conditions matter considerably. Oh. if you are given a plasmid, it is always a good idea to sequence it and make glycerol stocks. You can spend way too much time trying to optimize expression from a non-functional mutated plasmid. Good luck! Here is a list of the pET "system" (old style) https://research.fredhutch.org/content/dam/stripe/hahn/methods/biochem/pet.pdf
Hi there,
There is no golden rule for protein expression. Only experiment will tell you at the end.
In my experience, if the protein is small like 30 kDa then it is easy to recombinantly express and purify. If it is about 70 kDa it might be moderately difficult to express and purify.
I would AlphaFold the protein sequence with google collab servers and if you see a lot of helices and beta sheets then it probably expresses well. If you see a lot of coils and loops then it probably won't express well.
BL21(DE3) is standard E.coli cell line for protein expression. Arctic Express cells are good option if you are struggling. They coexpress a chaperone protein.
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