Clearly the ionic strength of a medium can affect N-H chemical shifts. One would expect changes in the HSQC spectrum from the non salt situation to one where salt is added. At a certain salt concentration the effect should asymptote to a final value.

and of course you will not be able to do this with a regular NMR tube and probe as the ionic strength is too high for a normal probe, shaped tubes and micro tubes can help I have recorded upto 0.5M in a micro tube with a microprobe before but beyond that I couldn't say, for a normal probe much above 150 mM you will get considerable signal loss and trouble matching the probe 

Thank you for your comments.

I have recorded 15N-1H HSQC of my protein in different salt concentration (from 0 M to 2 M NH4Cl) at pH=7.5, Temp=20 oC, using 3 mm NORELL nmrtube on Agilent DD2 700 MHz spectrometer equipped with an HFCN cold probe (Used DSS as an external reference).

I do observe one of glycine peak at 8.9256 ppm in 0M salt and it get upfield shift at 8.6088 ppm in 2 M salt (Difference is 0.3168 ppm) . What I am worried is that real or something wrong with my processing and referencing (Data are processed using NMRPipe) . 

I would highly appreciate your comments or any reference where HSQC has been recorded in high salt.

Thanking you,

Best Regards,

Anil

I don't think you should be worried about your referencing or processing if it's a single residue's amide shift that changed. You have many internal controls!

How large (and ordered) is this protein? Is the peak shifting in the fast exchange limit? It's probably just a minor change in the hydrogen bonding -- at higher salts there's less bulk water available to hydrogen bond to the Gly. So I guess I'm predicting that the Gly is solvent exposed (although that's not much of a prediction.) It's hard to invoke a more interesting conformational transition that is only reported by a single Gly!

Parenthetically, I disagree with Prof. Naider; I don't think protein amide chemical shifts show much of an ionic strength dependency unless there are concomitant changes in conformation (or hydrogen bonding as I'm speculating above.) Maybe he has a reference? Most proteins are more monodisperse in solutions with biologically plausible amounts of electrolytes, so I think most examples of 'salt-induced chemical shifts' reflect some intrinsic protein behavior. A good reference that uses model peptides should resolve this.

EDIT: Anil requests a reference where HSQCs are acquired in high salt. I've linked to a paper with HSQCs acquired in 5M GdmHCl.

http://www.ncbi.nlm.nih.gov/pmc/articles/PMC1134101/

To be exhaustive, it wouldn't be impossible for there to be a specific ion effect, e.g a weak protein binding site for the ammonium or (less likely) chloride ion. There could also be indirect salt effects on the pKa of a neighbouring residue, leading to a reorganisation of H-bonds. Maybe there is a millisecond dynamic exchange event at the Gly residue and the salt changes the thermodynamic balance between the states in exchange. All sorts of reasons are possible for a localised effect... 

Perhaps Ryan has a point. If you are finding only a change in one residue that is affected it does not appear to be a general ionic strength dependency.  I, however, find that to be strange, especially with ammonium chloride which is the salt of a strong acid and a weak base. I do not know what Ryan means by plausible amounts of electrolytes carried with the protein. If this means bound salts then their concentrations are unlikely to be comparable to those of the added ammonium chloride. By the way how do you buffer at pH7.5 in changing ammonium chloride concentrations. and do you measure the pH? Finally, it would seem likely that there will be a difference between buried amides and those exposed on the surface, as in non structured regions (disordered, irregular) of the protein. Buried backbone amides may not be sensitive to ionic strength but those that are exposed - I am not sure that would be the case.