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I have recorded 2D half filtered NOE experiments on a protein-protein complex and got very poor quality data, why would this be?

The interaction is in intermediate exchange, so presumably if my labelled protein is saturated to give me sharp peaks the partner will have begun to broaden out. Will the observed NOE suffer the same broadening or will it simply carry the chemical shift of its bound state? I thought it would also broaden but I am unsure of this.

Any help would be appreciated


edit: My assumption was that broadened resonances due to exchange in one protein would cause the intermolecular NOE to broaded also, what I was unsure about is whether I was missing somthing and say, in reality because the intermolecular NOE was originating only from the bound state, i.e. one of the exchanging species, whether it would still be broad or maintain the chemical shift of the bound species from which the NOE is coming from

asked Jul 12 '10 at 10:53

TomgA20's gravatar image


updated Jul 15 '10 at 08:17

Evgeny%20Fadeev's gravatar image

Evgeny Fadeev

could you clarify in what way the data is poor? - Evgeny Fadeev (Jul 13 '10 at 07:45)

Tom, I've merged your follow up into the question. This forum works a little differently from the traditional discussion groups. Q&A format is a lot easier to follow than threads. So if you need to follow up - either post a new question, edit an existing one, or post a comment. - Evgeny Fadeev (Jul 15 '10 at 08:20)

Normally people cannot edit other's posts, but I can because I moderate this site. - Evgeny Fadeev (Jul 15 '10 at 08:21)

Do you see two states in the spectrum? - Evgeny Fadeev (Jul 15 '10 at 08:27)

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Well maybe someone will correct me, but I'd think that you will see broadened NOESY peaks in the case of intermediate rate of exchange, regardless of using the isotope filter in the experiment.

If exchange rate is fast you may see transfer NOE's - correlations between resonances of unbound and bound species.

Complexes might have additional exchange "dimensions" in the multiple docking conformations which may contribute to more line broadening. For example aromatic groups can enhance broadening because the ring current chemical shift strongly depends on the orientation of the aromatic residue (I know of a case where replacing the aromatic residue with non-aromatic improved quality of the spectrum).


answered Jul 13 '10 at 07:56

Evgeny%20Fadeev's gravatar image

Evgeny Fadeev

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