Hi Romel,
The nanosecond motions you are seeing are interesting. Whether or not
they are real is a valid question. However like most NMR
spectroscopists, you are completely wrong in your assumption ;) You
just need to carefully read the second of the Lipari and Szabo papers,
something many people do not do, wherein they check the validity of
internal motions slower than the global tumbling of the molecule.
You can also perform your own tests by setting up synthetic models in
relax - just load a PDB file, set a diffusion tensor to the values you
like, set the internal model-free parameters, and then back calculate
the relaxation data. Then perform a simple single model model-free
fit and see what happens. I.e. what they do in the second Lipari and
Szabo paper (well, without relax). You will see that with no
experimental noise, you can easily extract internal motions slower
than the global correlation time. Think of it as follows: After one
time period of the global tumbling tau_m, the exponential total
correlation function has not hit zero! It takes a few periods of
tau_m before statistical zero is reached. Therefore other exponential
functions can be present in the auto-correlation function. In the
case of ellipsoidal diffusion, there are 5 exponential functions mixed
in together. So why can, for example, the isotropic diffusion with a
single exponential not be mixed in with another exponential coming
from the internal motions rather than the global tumbling process?
The answer, which is present in the original Lipari and Szabo papers,
is that it most definitely can. The problem of parameter extraction
is experimental, not theoretical! As soon as you add noise to the
synthetic relaxation data, it becomes harder and harder to extract the
slow internal motions reliably. Monte Carlo simulations can show this
too. Again this can be tested by adding white noise to the back
calculated relaxation data. I highly recommend you perform these
tests yourself so that you get a good idea of what you can obtain - it
will help you understand your system better. You can even use your
protein system for the synthetic data tests. Note that if the two
correlation times are identical, you will have problems. Just test
yourself and see what happens! By learning from these tests, you will
be able to formulate much stronger conclusions from your experimental
results.
As for your experimental data, another check would be to see if these
motions are present in the local tm models. I think I discussed this
in my second 2008 paper
(http://www.nmr-relax.com/refs.html#dAuvergneGooley08b or
http://dx.doi.org/10.1007/s10858-007-9213-3). You also have to
carefully think if a single diffusion tensor is valid for the entirety
of your system, an assumption which may not hold.
One question I have though is how did you change the model selection
protocol in the dauvergne_protocol.py auto-analysis? This is
hardcoded into relax, and the use of both AICc and BIC is invalid for
the searching of the universal solution (see
http://thread.gmane.org/gmane.science.nmr.relax.user/1356/focus=1357
and my 2007 paper at
http://www.nmr-relax.com/refs.html#dAuvergneGooley07 or
http://dx.doi.org/10.1039/b702202f). You can only use model selection
techniques where the fundamental derivation is based on the
Kullback-Leibler discrepancy
(http://en.wikipedia.org/wiki/Kullback%E2%80%93Leibler_divergence) or
the Fisher information metric. AIC and ICOMP model selection are
suitable, for example, but AICc and BIC are not as the first is a
deliberate divergence from the KL discrepancy and the second arises
from quite different Bayesian assumptions. To use AICc or BIC, the
universal solution equation of my 2007 paper would need to be
reformulated to match the very different fundamental concepts behind
these techniques and the model-free protocol redesigned to fit.
Regards,
Edward
P. S. Because of the number of bugfixes since relax 2.0.0, I would
recommend updating to relax 2.1.2
(http://marc.info/?l=relax-announce&m=135070664825024).
On 11 December 2012 10:28, Romel Bobby <rbob002@xxxxxxxxxxxxxxxxx> wrote:
Dear Ed and relax users,
I have recently used relax (ver 2.0.0) to obtain the model-free dynamics
of
a small protein with a molecular mass of 6 kDa. In fact, it's a protein
in
complex with a small peptide, but only the protein is 15N isotope
labelled.
R1, R2 experiments were recorded at 600, 800 and 900 MHz, and the
steady-state NOE at 800 and 900 MHz. Temperature calibration was
performed
before running the experiments for all the fields.
I used the fully automated analysis (dauvergne_protocol.py) and that
went
well. At the end of the run, the selected diffusion tensor had the form
of
an oblate spheroid with a global rotational correlation time of 3.2 ns.
Now, I have observed that some residues display slow internal
correlation
times (ts) that are in the range of 1-4 ns. Considering the tm is only
3.2
ns, this would mean, that motions slower than the overall tumbling were
captured. However, my understanding is and please correct me if I'm
wrong,
that except for Rex contributions, relaxation measurements are
insensitive
to motions on a time scale equal to or slower than the overall tumbling.
I'm a bit puzzled about the validity of these motions, that is to say,
are
they physically meaningful?
Is it possible that some of the motional models were not adequately fit
and
poorly chosen? Also, I've used the AICc selection method to reduce the
probability of overfitting.
Cheers,
Romel Bobby
Biomolecular NMR Research Group
School of Chemical Sciences/School of Biological Sciences
The University of Auckland
+64 (09) 3737599 Ext 83157
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Re: internal motions comparable to tm