Dear scientists
I do have a mammalian protein which i want to express in E.coli.as we know one of the important factors in protein expression is the mRNA secondary structure. It's better for the first 10 codons to be in a loop to be sure that this protein will be expressed well in E.coli. I have optimized my sequence ab lot and the best mRNA structure it gave me is this. I wonder if this can be expressed in E.coli?
Dear Nesa.
Rna structure or just one factor that can affect mammalian protein expression in e.coli. you have also to take in account the number of rare codons, presence of ptm as s-s bridge, presence of tm regions, e.coli strain user for the expression, domain expressed and addittion of an n- terminal tag. Thereore is really difficult to answer to yuor question. However when you di this kiñd of analisys you have to include also the region of the plasmid that contain the promoter and rbs. In fact The rna harpin prevent ribosome binding and the inhitation of the translation. Therefore the optimization need to be done in order to minimize interaction in this region. Sometimes the simplest solution is use an n-terminal tag ad gst, gb1, mbp, that is known to guarantee higher expression in e.coli. However this not mean that you will obtain a soluble and stable protein.
Good luck
Manuele
If you want a meaningful answer from an mRNA structure prediction, you need to include the 5' UTR as well (it looks from here as if you used only the protein-coding part of the mRNA).
Have you used the RNA calculator at https://www.denovodna.com/software/ ? It has a reverse-engineering mode that will provide the answer you are after.
Hope it helps!
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