Yes, you can separate by electrophoresis, that's why we use it, but we
cannot calculate accurate mass of complexes.
Maia
Jürgen Bosch wrote:
> Not quite correct, look into Blue Native PAGE. There you can seperate
> natively by mass.
>
> Jürgen
>
> ......................
> Jürgen Bosch
> Johns Hopkins Bloomberg School of Public Health
> Department of Biochemistry & Molecular Biology
> Johns Hopkins Malaria Research Institute
> 615 North Wolfe Street, W8708
> Baltimore, MD 21205
> Phone: +1-410-614-4742
> Lab: +1-410-614-4894
> Fax: +1-410-955-3655
> http://web.mac.com/bosch_lab/
>
> On May 19, 2010, at 1:31, Maia Cherney <[log in to unmask]> wrote:
>
>> Dear Jacob, I offer you my opinion.
>> Are you talking about electrophoresis? As far as I know it does not work
>> for the mass. The velocity of a protein depends on the charge at a
>> particular pH, the mass and shape of molecules etc. It's very difficult
>> to take all these things into consideration. Otherwise this would be a
>> very convenient method, much easier than the analytical centrifugation
>> or gel-filtration that are usually used. However, electrophoresis does
>> not work for mass determination. Besides, complex formation hugely
>> depends on the protein concentration. If you dilute your mixture, your
>> complexes might dissociate. There is equilibrium constant between
>> different types of complexes.
>>
>> Maia
>>
>>
>> Jacob Keller wrote:
>>> Dear Crystallographers,
>>>
>>> I am trying to optimize a native gel experiment of a two-protein
>>> complex, running the smallest-detectable amount of protein component A
>>> with varying amounts of component B.
>>>
>>> MW Charge MW/Charge
>>> A 22 -5 -4308
>>> B 17 -24 -702
>>>
>>> This experiment is partly to determine stoichiometry, but also to
>>> determine roughly the strength of the interaction.
>>>
>>> B definitely runs much faster than A alone, as predicted, but I am
>>> wondering what to expect with various oligomers. Should ABB run faster
>>> or slower than AB? What about AABB? Theoretically, AA should certainly
>>> run slower than A, and BB slower than B, simply because the
>>> mass/charge ratio is the same, but the overall mass is greater. But
>>> what happens when you have AAB, for example? There must be an equation
>>> relating the mass/charge and mass (and perhaps gel percentage) to the
>>> speed traveled in the gel--but what is the equation?
>>>
>>> Thanks for your consideration,
>>>
>>> Jacob
>>>
>>> *******************************************
>>> Jacob Pearson Keller
>>> Northwestern University
>>> Medical Scientist Training Program
>>> Dallos Laboratory
>>> F. Searle 1-240
>>> 2240 Campus Drive
>>> Evanston IL 60208
>>> lab: 847.491.2438
>>> cel: 773.608.9185
>>> email: [log in to unmask]
>>> *******************************************
>>>
>>>
>
>
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