*** For details on how to be removed from this list visit the ***
*** CCP4 home page http://www.ccp4.ac.uk ***
On Jan 10, 2007, at 17:39, Flip Hoedemaeker wrote:
*** For details on how to be removed from this list visit the ***
*** CCP4 home page http://www.ccp4.ac.uk ***
James:
Doesn't this ignore the use of B-factor restraints in the refinement
programs? Because the B-factors are in most cases refined in this
way,
modelling an atom in a position not supported by experimental
evidence, and
allowing its B-factor to inflate, will artificially inflate others
in the
residue as well, and correspondingly, will restrain its B-factor to
a value
lower than it should be.
Well, yes! B-values are restrained for a reason, being that the
atoms have
covalent bonds! The crystallographers choice of (lowering) the
weight on
this restraint is in the end an interpretation of the data, just like
leaving out atoms that are really there. Models are always wrong of
course,
the debate is what is more wrong and what is less wrong...
In the end how does one _really_ know the atoms are there?
Carboxylates may get decarboxylated by radiation damage, the sequence
could have been wrong etc. I find there is a significant difference
between modelling something into electron density or outside it.
James:
Furthermore, this practice would be expected to greatly increase
user error
because there is no clear indication of the atoms that were positioned
arbitrarily according to non-experimental terms. Hopefully the end-
user
will look at the model in the context of the structure factor data,
but as
we know this is not always done, and even worse, can't be done
where the
structure factors haven't been deposited. In this worse case
scenario, how
can someone be sure that any surface interactions between
sidechains with
middle-to-high B-factor is supported at all by the electron density?
Hopefully the crystallographer who models missing atoms in this way is
vigilant about depositing their structure factors!
A measured reflection contains contributions of all atoms in the
structure,
including the bulk solvent. The bulk solvent molecules are also not
visualized in the final model, but are taken into account during
refinement
anyway. If you leave atoms out of the refinement, you introduce or
at least
spread out errors on the remaining atoms. Not depositing structure
factors
is definitely a nono, I agree. Maybe we should also publish all the
restraints etc in the remark section as well, but that certainly
won't be
read by the majority of users!
Would you also not introduce errors by including in the refinement
atoms whose positions are not well determined by the data (i.e.
disordered)? Taking something into account (such as bulk solvent)
doesn't add parameters.
James:
I think building atoms in places not supported by the data without
at least
an occ=0 is the wrong approach. There are plenty of modelling
programs out
there that can do that for you, even humble SwissPDB viewer will
arbitrarily
reconstruct missing side chains and colour them differently to the
rest of
the protein.
The atoms are placed based on experimental evidence (neighboring
atoms),
plus geometry restraints (including rotamer statistics). Again,
model error
can only be assesed for the model as a whole, not just for
individual atoms
I think.
I think there is a clear difference between say a missing CG in a
threonine when you see everything else and it's very clear where the
missing atom has to be and a lysine where you only see CB and the
rest could be where ever. In the first case it's justified to let the
B-factor of the missing atom refine to >100 (which it probably
wouldn't do because we restrain it...) because it's reasonable to
assume that we have the equilibrium position right. After all B-
factors are supposed to describe harmonic motion about the
equilibrium position, and in the lysine case those would be entirely
arbitrary and the model would not provide much insight into where the
atoms really are...
Esko
Esko Oksanen, M.Sc., researcher
Macromolecular Structures Group
Research Program in Structural Biology and Biophysics
Institute of Biotechnology, University of Helsinki
Viikinkaari 1 P.O. Box 65
FIN-00014 Helsinki
FINLAND
tel. +358-9-19158903 fax +358-9-19159940
mob. +358-50-3771414
Skype ejoksane
[EMAIL PROTECTED]
Flip
--
Dr. James Irving
NH&MRC C.J. Martin Fellow
Division of Structural Biology
Wellcome Trust Centre for Human Genetics
Oxford University
Roosevelt Drive,
Oxford OX3 7BN
UK
email: [EMAIL PROTECTED]
phone: +44 1865 287 550
