Is there any Gram positive strain that could be used for over expression of membrane protein complexes (like the gram negative BL21 cells)?
Most of the time, when you wish to purify and crystallize a protein or a complex, the method of choice is to transform E.coli BL21 or Rosetta with the corresponding plasmid(s).
However, some complexes may not exist or cannot be stable in these strains, but only in Gram positive bacteria.
Could anyone provide any suggestion/reference/help on this topic?
Cheers.
Hi Patrice
Maybe this helps for your search:
http://www.nature.com/nrmicro/journal/v12/n3/fig_tab/nrmicro3199_F2.html
Be carefull about the pathogenity of your new host strain....
I have good experience with yeast expression, if activity or stability is problem, or specially for Membrane-based Proteins with Pseudomonas stutzeri, this MO is really interesting for transformation problems:
http://jb.asm.org/content/153/1/93.abstract
if you want to try a safe and simple solution with a traditional lab-Coli, try Stratagenes
JM101.
This is a very old strain that is not used for cloning today, but it has a few interesting and unique features. The stain grows at 30°C with the same rates like standard colis at 37°C, so you can reduce the temperature in ypur expression and have less termal stress for proteins. The strain has a very high permeable membrane, so proteins located there have a better activity than other strain.
The reason for this I think is production of enormous amounts of RNA, far too much for a regular cloning coli (you find it in the supernatant).
This strain is used for succesfull technical fermentation of some membrane-based proteins, that dont show activity in other Host-sytems, also not in expression optimized BL21s.
It is a little sensitiv to temparature, so stay at 30°C for cultivation.
good luck
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