Help protein localization

Frank O. Fackelmayer Frank at Fackelmayer.de
Wed Jul 10 09:16:43 EST 2002


I second Frank´s and Artem´s answers: experiments will show you where
your protein is!
A good starting point is to perform subcellular fractionation to
separate as many different organelles, and trace your protein by western
blotting those. For all organelles I know there exist typical marker
proteins (for many of which antibodies are commercially available) that
you can (and should!) use as controls for the fractionation.

In vivo localization of a protein can be visualized by gfp fusions just
as you did, but you should consider that GFP is quite big and might
interfere with your protein's function and/or localization.

Frank



deleonardis at farmbiol.uniba.it wrote:
> 
> Hi,
> 
> I'm a Ph d sudent and I'm working for an ipotetical mitochondrial or
> 
> peroxisome carrier protein (human)  (maybe Aprille's carrier) and I just
> 
> work with my protein fused with GFP, but I dont'have any evidence of
> 
> mitochondrial localization for my protein (i used CHO cell). So, the best
> 
> software for alignment give an high omology with Ca2+ binding solute carrier
> 
> protein  of rabbit.....but other softwares say that my protein is
> 
> cytosolic(Prosite, ecc) . I study  in vitro (ricostituited into
> 
> proteoliposome) the transport of my protein cloned and overexpressed in
> 
> E.coli.........
> 
> Does anyone help me, such as say me the best software for subcellular
> 
> loclization ....ecc.
> 
> Thanks.
> 
> http://www.biowww.net/index.php/forum/forumlist/1/



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