I'm running SDS-PAGE in reducing condition, but always my recombinant proteins migrate to the wrong size. Does anyone know why?
What means wrong size? Smaller than you get it from the normal environment?
If its much smaller, than you might have split a a multimeric protein into its monomers. If its a little smaller, than this might be come up to missing post-translational modifications.
Your protein may be partially degraded. Which kind of protease inhibitor do you use?
If it's a small difference, it may be a characteristic of the protein itself. Recombinant and synthetic human alpha-synuclein show up in gels at around 17 kDa, even though their molecular weight is 14.6 kDa. I can confirm this for recombinant aSyn, which we have purified in our lab and characterized via immunoblotting and mass spec. The immunoblots show a monomeric band around 17 kDa, while mass spec clearly shows the proper molecular weight of 14.6 kDa. I would recommend using some other method to check that your protein is the proper molecular weight, whether it be mass spec, analytical ultracentrifugation, SEC with appropriate standards, or another method.
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