Dear Nat,
yes, I fully agree - all these restraints that improve the geometry
either by restraining to high-resolution structures or by introducing
H-bond restraints for secondary structures are very useful for
low-resolution structures!
I see your argument with the Ramachandran plot. But imagine a set of
very strong non-bonding/bonding restraints that would result in an
absolutely clean Ramachandran plot for any structure, then the
Ramachandran plot would become useless even in the absence of any
phi/psi-restraints. So, I prefer to err a bit on the safe side here by
saying "not a truly independent measure".
Personally, I think, that ALL refinement programs, including the
real-space refinement in Coot, would benefit from inclusion of proper
H-bonding terms (something, that for instance the very old X-Plor
version did), since this would automatically restrain secondary
structures and other hydrophilic interactions to some reasonable
geometry, even at very low resolution.
Best regards,
Dirk.
Am 26.09.11 16:17, schrieb Nat Echols:
On Mon, Sep 26, 2011 at 1:53 AM, Dirk Kostrewa
<[email protected] <mailto:[email protected]>> wrote:
when I played with H-bond restraints for secondary structures for
the refinement of a 4.3 A structure (only a few weeks before they
were introduced in phenix), I've made the following observation:
at low resolution without H-bond restraints for secondary
structures, the carbonyl groups of these secondary structures take
the liberty within their globbish electron densities to deviate
from their ideal H-bond conformation, resulting in a tight "belt"
of outliers around the preferred Ramachandran regions, with
typical deviations of only a few degrees. Introducing the
additional H-bond restraints for maintaining secondary structures
pulls these outlier carbonyl groups back into the preferred
Ramachandran regions. In my case, the number of Ramachandran
outliers was reduced to less than one half! Although, these H-bond
restraints do not directly include information about allowed
Ramachandran regions, the Ramachandran plot is actually affected
by these restraints. Thus, at least in my opinion, the
Ramachandran plot is then not a truly independent measure for
model quality, anymore. The same holds true for all geometrical
restraints, of course.
It depends on how strictly you assess the "independence" of validation
criteria. The Ramachandran plot is considered valid in most cases
because refinement programs traditionally do not restrain phi and psi
angles, so we need to rely on the accuracy of the data (and our
placement of atoms) and various complementary geometry restraints
(especially nonbonded) to keep residues in the favorable regions of
the plot. There are a variety of ways to make the plot better by
modification of the model and/or restraints (adding hydrogens,
increasing the weight on the nonbonded restraints, secondary structure
restraints, etc.), none of which are as drastic as directly
restraining the model to the plot. I don't really view this as
biasing the plot, for two reasons: a) the quantity being measured is
independent of the quantity restrained, and b) at least in my hands,
these modifications never completely fix the problem of Ramachandran
outliers. (It's the loop regions that are really awful.)
Anyway, I don't think anyone should feel bad about using this kind of
restraint at low resolution. The caveat is that of all the
specialized restraints that we (Jeff Headd and I) have been testing
for low-resolution refinement (in Phenix), nothing works nearly as
well in preserving good geometry, and usually improving the R-factors,
as restraining model parameters to a related high-resolution
structure, when one is available. Fortunately, every modern
refinement program has this ability in some form, and I expect that
this is going to have the most impact in improving the overall quality
of low-resolution structures.
-Nat
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Dirk Kostrewa
Gene Center Munich, A5.07
Department of Biochemistry
Ludwig-Maximilians-Universität München
Feodor-Lynen-Str. 25
D-81377 Munich
Germany
Phone: +49-89-2180-76845
Fax: +49-89-2180-76999
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