That's interesting. Thanks.
Maia
Nadir T. Mrabet wrote:
> Maia speaks about native PAGE for which protein mobility (migration)
> depends on 3 different parameters as she states: charge, mass and shape.
> Blue native PAGE, which might be the answer to Jacob's question, is a
> 2D gel: Native in the first direction, then SDS-PAGE in the second one.
> You actually need both data to infer stoechiometry and subunit
> composition.
>
> Nadir
>
> Pr. Nadir T. Mrabet
> Structural& Molecular Biochemistry
> Nutrigenex - INSERM U-954
> Nancy University, School of Medicine
> 9, Avenue de la Foret de Haye, BP 184
> 54505 Vandoeuvre-les-Nancy Cedex
> France
> Phone: +33 (0)3.83.68.32.73
> Fax: +33 (0)3.83.68.32.79
> E-mail: Nadir.Mrabet<at> medecine.uhp-nancy.fr
>
>
>
> On 19/05/2010 13:01, 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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