fusing several gfp's to a protein

Frank O. Fackelmayer Frank.Fackelmayer at uni-konstanz.de
Wed Jan 26 03:53:14 EST 2000


Raavo Toomela wrote:

> hi,
> 1. what problems may arise, when fusing 2 (at least) gfp's to a coat protein
> in size about 30 kdaltons.

a. you may have plasmid stability problems in E.coli that does not tolerate
long stretches of repeated DNA on the same plasmid.
b. the tags may interfere with the protein´s natural function, because GFP is
comparable in size to your protein. That´s like dressing up with a coat that
has your weight (or multiples thereof), and expecting that you move as you
would normally...
WHY would you like to add more than 1 tag?

>
> 2. how to avoid diffusion of gfp-protein complex in cells? size must be over
> 50 kDa?
>

GFP fusion proteins are usually made to investigate proteins in vivo. So, if
your protein diffuses throughout the cell, the fusion should also do that (or
you cannot learn anything significant about your protein).

Frank






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