recombinant protein

Malcolm M. Campbellcampbell campbell at cict.fr
Fri Feb 5 07:11:53 EST 1993


Fellow netters,

I realize that the following question is becoming a FAQ (and I'm kicking 
myself for not saving previous postings) but...

What is the consensus out there regarding the best recombinant protein 
expression system based on the following criteria:

1) system must produce a functional protein (ie. with catalytic activity);
2) protein is easily purified to _homogeneity_ AND in high quantities 
   (think mg's), and;
3) the cells carrying the recombinant protein are easily transformable 
   and can be grown using standard bacterial culture facilities?

We have already expressed active protein (from the cDNA encoding our plant-
derived protein) using the pT77 system - it works great if we want to assay 
catalytic activity in crude preps.  However, now we want pure protein for 
NMR analysis of protein/substrate/inhibitor interactions and, of course, 
crude preps just aren't going to cut it.  After having developed a number 
of protein purification protocols for this enzyme from a number of plant 
species, we aren't too enthuastic about having to tackle the same problems 
with our bugs.  A nice, simple protocol would be great...

Can anyone out there help us?

Thanks in advance.

Cheers,

Malcolm
++++++++++++++++++++++++++++++++++++++++++++++++++++++++++++++++++++++
Malcolm M. Campbell                          + phone: (33) 61 55 69 05 
Equipe Lignines                              + FAX:   (33) 61 55 62 10
Centre de Biologie et Physiologie Vegetales  + internet:
URA CNRS No. 1457                            + campbell at cix.cict.fr
Universite Paul Sabatier                     +
118 route de Narbonne                        +
31062 Toulouse cedex                         +
FRANCE                                       +
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