His tagged protein expression

Matthew Hicks baps9 at central.susx.ac.uk
Thu Aug 6 18:23:58 EST 1998


If the protein is non-covalently linked as a trimer the SDS will (almost
certainly!) denature it - ie the protein will run as monomer on the gel.
However, if the polypeptide chains are disulphide linked then you will
have to reduce these (covalent) bonds to observe the monomer molecular
weight on your gel:  
To reduce the disulphide bonds the best plan is to use a mercaptan (eg.
mercaptoethanol: HSCH2CH2OH - smells like farts/eggs though! ) in your
gel.   These are usually used in "denaturing" gels - check your recipie.
If there is a disulphide-reducing agent in your gel then some weird
stuff is going on (have you checked to see if the nucleotide sequence
has been inserted more than once, ie the protein is being expressed as
three repeats of the sequence, - sorry I am not a molecular biologist
so I may be talking out of my ****/patronising you - but cannot think
what else may be happening!).
Let me know how you get on and good luck.

Matt Hicks
University of Sussex
Falmer
East Sussex
UK




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