One of my main concerns is that the unfolded protein itself would irreversibly aggregate and then wouldn’t interact with the folded protein.
I think DSC (differential scanning calorimetry) should be performed first to characterize the state of both proteins and their potential “complex”. By analyzing the DSC data, you should have an idea what kind of enthalpy change is involved during the complex
formation. If you see the unfolded protein does become folded, then ITC experiments are worth trying. A combination of ITC/DSC experiments may help you tweak out the exact interaction parameters.
Wendy
--------------------------------------------------
Wendy Yang
Manager, Biocalorimetry Laboratory
Center for Biophysical Sciences & Engineering
Tel: 205-975-2450
Fax: 205-934-0480
Email:
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From: CCP4 bulletin board [mailto:[log in to unmask]]
On Behalf Of Anita P
Sent: Friday, March 14, 2014 8:57 AM
To: [log in to unmask]
Subject: Re: [ccp4bb] ITC with unfolded proteins
That is a very interesting question, which I would request the seniors out there to give their insights on.
I was imagining that a recombinant purification of an unfolded partner would aggregate which would cause trouble in ITC. Am I correct in this theory?
Would love to have more insights.
thanks in advance
Anita
On Fri, Mar 14, 2014 at 7:18 PM, <[log in to unmask]> wrote:
Hi,
I think the experiment is doable, but how would you decouple
protein-protein interaction from folding of the unfolded
protein due to protein interaction?
Reza
Reza Khayat, PhD
Assistant Professor
The
Department of Chemistry, MR-1135
Tel. (212) 650-6070
---- Original message ----
>Date: Fri, 14 Mar 2014 18:07:48 +0530
>From: CCP4 bulletin board <[log in to unmask]> (on behalf
of Anita P <[log in to unmask]>)
>Subject: [ccp4bb] ITC with unfolded proteins
>To: [log in to unmask]
>
> Hello everyone,
> I have a query for the scientists working on
> protein-protein interaction.
> It is known that some proteins exist in unfolded or
> molten globule state and attain structure on
> interaction with other folded proteins.
> Many a times, it is difficult to obtain the
> structure of these complexes.
> Is it possible to quantitatively determine the
> thermodynamics of interaction between an unfolded
> protein and a folded protein using ITC? Later may be
> perform an alascan to determine the residues of the
> unfolded partner involved in the interaction.
> Please share your ideas
> cheers**
> Anita