Susanne and all,
I'll jump in late and offer a possible different interpretation.
Inferring masses from size-exclusion chromatography is always subject to
large errors---I have seen so many cases where a putative dimer is really a
monomer, and also a case where a protein said to be a monomer by SEC was
really a trimer. If you don't mind being off by factors of 2-3 either way,
SEC is fine.
So one interpretation would be that when your proteins are by themselves
they are in a very extended conformation (maybe even partially unfolded) and
so elute about where a dimer should elute. Then when you mix them together
they really do form a heterodimer but it is so much more globular (and/or
perhaps sticks more to the column) that it actually elutes later than the
monomers.
Overall, this is yet another example of why one needs rigorous, absolute
methods for determining solution mass like analytical centrifugation or
classical light scattering (and what keeps our company in business!)
John Philo
Director of Biophysical Chemistry
Alliance Protein Laboratories
www.ap-lab.com
"Susanne Rohrer" <rohrersusanne.spam at hotmail.com> wrote in message
news:3CD938B0.93F13889 at hotmail.com...
> > yes, it may well be that the protein form homodimers but not
> > heterodimers in absence of "something" else, but that would not explain
> > why the homodimers are split when the two proteins are mixed.
>> If this was an artefact, what could cause it...?
>> > Does one (or both) of the proteins have an enzymatic activity that could
> > modify the other protein? Idea: Protein A (x)ylates Protein B. Protein B
> > dimers form only in the un(x)ylated form, so they cannot be reformed
> > after the reaction. For BOTH your homodimers to split, it could be a
> > crosswise reaction: A-dimers meet B-dimers, (x)ylate each other and the
> > complexes break apart.
> > Sounds weird, but probably not too weird to occur in nature...
>> Yes that would be plausible if they had such an activity. But I don't
think
> they do. They are bacterial proteins involved in cell wall synthesis - It
> is known what they do (add an amino acid from a tRNA to the peptidoglycan)
> but no one has ever looked if they are themselves modified in a way.
>>
