I am working on a hyperthermophilic protein which is dimeric in its native state. I am checking at what guanidine concentration the dimer dissociates into a monomer. One way to do so is to employ static light scattering to deduce the molecular weight of the assembly. However, in the equation for static light scattering, we have one parameters dn/dc ( refractive index increment of the polymer under investigation). For a protein in aqueous buffer, we can easily consider it equal to 0.185 ml/g. I wonder how to calculate the dn/dc of a protein solution in a higher guanidine containing buffer. e.g sodium phosphate buffer with 4 M guanidine hydrochloride? Does it remain the same as in aqueous buffer?
Probably not. IF you know the concentraion of protein, it is esay to determine dn/dc by MALS-DRI. I am sure you can find procedure in manual. if you donot know the concentartion of protein. you can use 0.185 for relative comprision.
If you are concerned about dn/dc of your protein in your buffer, and you are really concerned that you need a high-precision answer, you might be better off measuring it directly for your protein in your buffer. Three or four points should be enough. I realize that you may have to hunt to find a working differential refractometer. (If anyone in North America wants a contact, I can supply it, but shipping biologicals around the world under modern conditions may be challenging. Please contact me via email and I will offer a contact. The lucky people with the old Bryce-Phoenix units should take good care of them.
Besides, measuring it yourself will silence an obvious annoying referee critique.
I agree with Georges: the best way is to measure it employing a short of differential refractometer (obviously calibrated!) and to get a perfect linear response in order to get a dn/dc with good enough precision for suing it for molecular weight determinations ...
As a perhaps-alternative, depending on exactly what you want to do and how the system behaves, you could look at I/c as a function of protein c. If the material is a monomer in guanidine at low protein concentration, and a dimer at larger, then you do not need to know dn/dc in order to see the c at which there is disassociation; that c might then depend on guanidine concentration in a way you could establish with a series of I/c curves. That is not the same as the approach you are considering, and may not work for your system. In particular, you may not be able to get adequate measurements at lower protein concentrations via lack of scattering intensity.
If you would like to investigate protein dimerization, you could easily use MicroScale Thermophoresis! MST is a powerful method to study molecular interactions in any buffer system, therefore also in GuaHCl or Urea. So you can directly compare the dissociation constant for your dimeric protein in pure buffer and in different concentrations of your denaturing agent.
Here is a nice publication where MST has been employed to study dimerization of a protein:
http://www.nanotemper-technologies.com/technology/publications/article/cell/
You need to be careful with this. Guanidine has a very high refractive index, and will dramatically affect dn/dc at high concentration. According to this http://www.sigmaaldrich.com/catalog/product/sigma/50937, the refractive index of 8M Guanidine HCl is 1.465, which is almost that of protein (i.e., dn/dc is almost zero). So at 4M dn/dc might be ~ 1/2 the usual value of 0.185. There is a dramatic change in dn/dc with that much extra stuff in the buffer, and it will impose a constant negative slope in apparent Mw vs. [GuHCl] if the protein's MW and concentration are fixed.
However, all is not lost. You should see a distinct kink or 'knee' in the curve of MW vs [GuHCl] where the dimer dissociates, if you take a closely - spaced concentration series. Good luck!
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