I work with a protein that apparently likes acidic pH and I suspect that it do change its structural conformational when I change the pH. Apart from crystallography, NMR, circular dichroism, gel filtration and analytical centrifugation, anyone could suggest an alternative experimental method to confirm or at least have more evidence to point out that my protein is actually changing its conformation?
Thanks in advance and be safe with this coronavirus issue!
Sorry I forgot to indicate that I also tried Differential scanning fluorimetry
A powerful technique for studying conformation of polymers in general, in particular in their native environments, is small-angle x-ray scattering (SAXS). To study the very small scattering angles, a long beam path length is typically required, so the tool requires a large vacuum tube and a lot of lab floor space. It isn't a typical bench top tool. There are several requirements for the sample preparation, such as dilution and collection of an appropriate 'background' signal. The analysis is also non-trivial. But I think it is a great tool, in particular to directly study the shape at various pH.
Another technique you can look into, which is perhaps a less expensive tool to find, is dynamic light scattering (DLS). I believe that technique can't give you as much detailed information about the shape of the molecules, but rather their hydrodynamic volumes. But I'm not an expert in that technique. Good luck.
I know FTIR can help to resolve changes in higher order structures of proteins. We used it for BSA (and gelatin) before and after cryomilling to see if anything had changed.
Though, I'm not an expert and FTIR. But it might be worthwhile to check it.
Good luck
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