low level expression vectors

Koen A.L. De Smet k.desmet at ic.ac.uk
Fri Apr 11 05:53:00 EST 1997


I have used the His-tagging vector pQE (Qiagen) for the overexpression 
and purification of several recombinant mycobacterial proteins. In most 
cases, the proteins were insoluble. 
I then made my own His-tagging vector for expression in mycobacteria, 
and have now found that 4 out of 4 proteins are soluble in this system. 
The level of expresion is still fairly low (say 10-20 x more than from 
the WT gene), but at least I have functional protein. 
There are two explanations. Either these proteins prefer being expressed 
in mycobacteria, or the overexpression makes them insoluble in E.coli.
To test this, I could try and put the genes in E.coli with a promotor 
that provides low expression. Does anybody know if there are such 
vectors commercially available? Ideally, I would want to use one with a 
His-tag.


_______________________________________________________________

        Dr Koen A.L. De Smet
        Research Fellow
        Department of Medical Microbiology
        Imperial College Medical School at St Mary's
        Norfolk Place
        London W2 1PG
        Great Britain
                                     
        Tel: (+44)-(0)171-594 3946  
        Fax: (+44)-(0)171-262-6299   
        Email: k.desmet at sm.ic.ac.uk
	       http://www.sm.ic.ac.uk/medmicro/home.htm
_______________________________________________________________

"If we knew what it was we were doing,
 it would not be called research, would it?".   --   A.Einstein
_______________________________________________________________



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