Re: [ccp4bb] secondary structure restraints [was: Phenix version 1.6.1 released]
On Wed, Mar 31, 2010 at 8:47 AM, Dirk Kostrewa kostr...@genzentrum.lmu.dewrote: this is very interesting! From the list of changes, it appears that in version 1.6.1, you use a similar idea to implement hydrogen bonds via DSSP during refinement that I used to stabilize the 4.3 A refinement of Pol II in complex with TFIIB [1]: . . . Could you please give more details of how this is implemented? What are your hydrogen bond target distances and sigmas? Do you update this list during refinement? We're still testing this method (thus alpha version in Paul's email); my initial experiments indicate that it may improve R-free by up to 0.5% at moderate resolution, and generally keeps R-work and R-free closer than would otherwise be the case. It can also make R-free worse, although I think this happens less frequently. I had expected this to be resolution-dependent, but I tried it on partially built structures at either 2.25A and 3.1A and the results looked similar. Part of the problem is deciding how to filter outliers; the default behavior is to throw out any bonds greater than a specified threshold (e.g. 2.5A for H-O). KSDSSP (UCSF's free clone) has some quirks with respect to helix assignment which result in incorrect bonding if you take the results as-is without filtering. However, for poor models at low resolution, leaving the outliers in may be essential. All of the restraint parameters are adjustable, but I found that 1.975A for H-O was appropriate, and I left the sigma at 0.05 but 0.02 is sometimes better. We don't update the bonds during refinement, but I suppose we could - haven't done enough testing yet to know whether this is helpful. I used a simple list of additional 2.9 A target-bond-distances between N and O with a target sigma of 0.05 A. This list was determined with DSSP and a self-made Fortran95 program using a user-defined energy-threshold prior to refinement and was kept constant during refinement. Currently PHENIX uses a 3.0A distance for N-O (sigma also 0.05), but that wasn't rigorously derived. My gut feeling is that N-O bonds are problematic, partly because the distribution of distances (in the Richardson lab's Top500 database) appears to be bimodal - I think this results from the difference between parallel and anti-parallel sheets. We probably need to account for this when calculating the bonds. I also found that the restraints usually improved R-free more when hydrogens were used, but I haven't tried anything close to a 4.3A structure yet. At that resolution, anything that keeps the helices wound probably helps. I hadn't realized that you used similar restraints in the PolII/TFIIB paper - I'd be interested in knowing more about this, since I haven't found many useful references so far. (I couldn't find the page on the BUSTER wiki, by the way.) Personally, I think, using secondary structure hydrogen bonds should be an option in every refinement program, especially at lower resolution!!! The BUSTER Wiki describes the procedure that I used. For REFMAC, I haven't seen anything similar, yet. If you don't mind using PHENIX for this: phenix.secondary_structure_restraints model.pdb format=refmac I haven't made it as far as testing the output with REFMAC, but it's there if anyone wants to try. FYI, the program is still clumsy about switching to N-O bonds when hydrogens are absent, so the argument h_bond_restraints.substitute_n_for_h=True may be necessary. -Nat
Re: [ccp4bb] Secondary structure restraints
to my knowledge, none of the existing reciprocal-space refinement programs is really suitable for low-resolution refinement. In my For what it's worth, I've had good luck with refmac5D, which incorporates an SAS target into model refinement (I believe this was has now been incorporated into the mainstream refmac branch semi-recently). Making full use of the chemistry/geometry information is important, but so is using as many observations as you can get your hands on. Pete
Re: [ccp4bb] Secondary structure restraints
Dear Phil CCP4ers, to my knowledge, none of the existing reciprocal-space refinement programs is really suitable for low-resolution refinement. In my opinion, what is clearly missing, are automatic hydrogen-bond- restraints that would stabilize all secondary structures and other structurally important H-bonds during refinement at low resolution. The old X-Plor program had hydrogen bonds parametrized. In real space refinement, this is also missing in Coot that screws up structures at low resolution. I use Moloc in these cases, because it accounts for all hydrogen bonds and does stable real space refinement of proteins and nucleic acids at any low resolution. I really hope, that also the chemical knowledge of the structurally important hydrogen bonds gets incorporated both into reciprocal space and real space refinement programs in the future, since more and more biologically important structures are solved at medium to low resolution!!! Best regards, Dirk. Am 08.01.2009 um 18:09 schrieb Phil Evans: Does anyone have a good way of imposing secondary structure restraints in a low resolution refinement? I've done this in the past as hydrogen bond distance restraints within helices, input to refmac as LINKs , with the list generated with a little program and certain amount of pain refmac now accepts an explicit list of external restraints, as does phenix.refine, but I'm looking for a way of generating these lists for quite a large structure without too much hackery, perhaps from a hydrogen-bond or secondary structure assignment program. Helices are reasonably straightforward (I can see how to do them from eg DSSP), but sheets are more complicated. Any suggestions? I'm sure that someone must have done this Phil *** Dirk Kostrewa Gene Center, A 5.07 Ludwig-Maximilians-University Feodor-Lynen-Str. 25 81377 Munich Germany Phone: +49-89-2180-76845 Fax:+49-89-2180-76999 E-mail: kostr...@lmb.uni-muenchen.de ***
Re: [ccp4bb] Secondary structure restraints
Phil Evans wrote: Does anyone have a good way of imposing secondary structure restraints in a low resolution refinement? I've done this in the past as hydrogen bond distance restraints within helices, input to refmac as LINKs , with the list generated with a little program and certain amount of pain refmac now accepts an explicit list of external restraints, as does phenix.refine, but I'm looking for a way of generating these lists for quite a large structure without too much hackery, perhaps from a hydrogen-bond or secondary structure assignment program. Helices are reasonably straightforward (I can see how to do them from eg DSSP), but sheets are more complicated. Any suggestions? I'm sure that someone must have done this Phil It was in PROTIN! Should look back at that code.. E
[ccp4bb] Secondary structure restraints
Does anyone have a good way of imposing secondary structure restraints in a low resolution refinement? I've done this in the past as hydrogen bond distance restraints within helices, input to refmac as LINKs , with the list generated with a little program and certain amount of pain refmac now accepts an explicit list of external restraints, as does phenix.refine, but I'm looking for a way of generating these lists for quite a large structure without too much hackery, perhaps from a hydrogen-bond or secondary structure assignment program. Helices are reasonably straightforward (I can see how to do them from eg DSSP), but sheets are more complicated. Any suggestions? I'm sure that someone must have done this Phil
Re: [ccp4bb] Secondary structure restraints
XPLO2D from the USF-Suite does this: snippet from manual You feed it a PDB file of the model to which you want to restrain your refinement model (e.g., that high-resolution native structure you already have, even though it may be in a different spacegroup or with different domain orientations). The program generates an X-PLOR include file which contains DIHEdral statements for the PHI, PSI, CHI-1 and CHI-2 torsions of the protein. If you protein contains a hinge region, simply remove or comment-out the relevant PHI and PSI restraints. If you don't want to impose restraints on CHI-1 and/or CHI-2, set the corresponding weights to zero (at the top of the X-PLOR include file). I never tried, but this will be compatible with phenix as well. Cheers Eckhard Phil Evans schrieb: Does anyone have a good way of imposing secondary structure restraints in a low resolution refinement? I've done this in the past as hydrogen bond distance restraints within helices, input to refmac as LINKs , with the list generated with a little program and certain amount of pain refmac now accepts an explicit list of external restraints, as does phenix.refine, but I'm looking for a way of generating these lists for quite a large structure without too much hackery, perhaps from a hydrogen-bond or secondary structure assignment program. Helices are reasonably straightforward (I can see how to do them from eg DSSP), but sheets are more complicated. Any suggestions? I'm sure that someone must have done this Phil -- Eckhard Hofmann eckhard.hofm...@bph.ruhr-uni-bochum.de Ruhr-Uni Bochum AG Proteinkristallographie, LS Biophysik, ND04/316 44780 Bochum Tel: +49-(0)234/32-24463, Sekr. -24461, FAX: -14762
Re: [ccp4bb] Secondary structure restraints: Oops
Hi Phil, sorry, haven't read you question properly. No idea how to get easily from top/par to cif for refmac. Probably would need a little scripting, but that's been exactly your question ... Eckhard XPLO2D from the USF-Suite does this: snippet from manual You feed it a PDB file of the model to which you want to restrain your refinement model (e.g., that high-resolution native structure you already have, even though it may be in a different spacegroup or with different domain orientations). The program generates an X-PLOR include file which contains DIHEdral statements for the PHI, PSI, CHI-1 and CHI-2 torsions of the protein. If you protein contains a hinge region, simply remove or comment-out the relevant PHI and PSI restraints. If you don't want to impose restraints on CHI-1 and/or CHI-2, set the corresponding weights to zero (at the top of the X-PLOR include file). I never tried, but this will be compatible with phenix as well. Cheers Eckhard Phil Evans schrieb: Does anyone have a good way of imposing secondary structure restraints in a low resolution refinement? I've done this in the past as hydrogen bond distance restraints within helices, input to refmac as LINKs , with the list generated with a little program and certain amount of pain refmac now accepts an explicit list of external restraints, as does phenix.refine, but I'm looking for a way of generating these lists for quite a large structure without too much hackery, perhaps from a hydrogen-bond or secondary structure assignment program. Helices are reasonably straightforward (I can see how to do them from eg DSSP), but sheets are more complicated. Any suggestions? I'm sure that someone must have done this Phil -- Eckhard Hofmann eckhard.hofm...@bph.ruhr-uni-bochum.de Ruhr-Uni Bochum AG Proteinkristallographie, LS Biophysik, ND04/316 44780 Bochum Tel: +49-(0)234/32-24463, Sekr. -24461, FAX: -14762
Re: [ccp4bb] Secondary structure restraints: Oops
If top/par file could be converted to the following type of instructions then you do not need to define everything in cif file (these are for torsion angles, all other restraints can be defined similarly) General torsion angle restraints for any quartet of atoms: external torsion first chain [ch] residue [res] insertion [ins] atom [n] [altecode [a]] next chain [ch] residue [res] insertion [ins] atom [n] [altecode [a] ] [symm y/n] next chain [ch] residue [res] insertion [ins] atom [n] [altecode [a] ] next chain [ch] residue [res] insertion [ins] atom [n] [altecode [a] ] [symm y/n] value v sigma s period p Exampl external torsion first chain A residue 220 atom C next chain A residue 220 atom CA next chain A residue 220 atom C next chain A residue 221 atom N value -60 sigma 10 period 1 regards Garib On 8 Jan 2009, at 18:14, Eckhard Hofmann wrote: Hi Phil, sorry, haven't read you question properly. No idea how to get easily from top/par to cif for refmac. Probably would need a little scripting, but that's been exactly your question ... Eckhard XPLO2D from the USF-Suite does this: snippet from manual You feed it a PDB file of the model to which you want to restrain your refinement model (e.g., that high-resolution native structure you already have, even though it may be in a different spacegroup or with different domain orientations). The program generates an X-PLOR include file which contains DIHEdral statements for the PHI, PSI, CHI-1 and CHI-2 torsions of the protein. If you protein contains a hinge region, simply remove or comment-out the relevant PHI and PSI restraints. If you don't want to impose restraints on CHI-1 and/or CHI-2, set the corresponding weights to zero (at the top of the X-PLOR include file). I never tried, but this will be compatible with phenix as well. Cheers Eckhard Phil Evans schrieb: Does anyone have a good way of imposing secondary structure restraints in a low resolution refinement? I've done this in the past as hydrogen bond distance restraints within helices, input to refmac as LINKs , with the list generated with a little program and certain amount of pain refmac now accepts an explicit list of external restraints, as does phenix.refine, but I'm looking for a way of generating these lists for quite a large structure without too much hackery, perhaps from a hydrogen-bond or secondary structure assignment program. Helices are reasonably straightforward (I can see how to do them from eg DSSP), but sheets are more complicated. Any suggestions? I'm sure that someone must have done this Phil -- Eckhard Hofmann eckhard.hofm...@bph.ruhr-uni-bochum.de Ruhr-Uni Bochum AG Proteinkristallographie, LS Biophysik, ND04/316 44780 Bochum Tel: +49-(0)234/32-24463, Sekr. -24461, FAX: -14762
Re: [ccp4bb] Secondary structure restraints: Oops
I would guess that it would be easier to restrain a helix by hydrogen bond lengths rather than by phi/psi torsion angles, and that could work for sheets as well. Phil On 8 Jan 2009, at 19:05, Garib Murshudov wrote: If top/par file could be converted to the following type of instructions then you do not need to define everything in cif file (these are for torsion angles, all other restraints can be defined similarly) General torsion angle restraints for any quartet of atoms: external torsion first chain [ch] residue [res] insertion [ins] atom [n] [altecode [a]] next chain [ch] residue [res] insertion [ins] atom [n] [altecode [a] ] [symm y/n] next chain [ch] residue [res] insertion [ins] atom [n] [altecode [a] ] next chain [ch] residue [res] insertion [ins] atom [n] [altecode [a] ] [symm y/n] value v sigma s period p Exampl external torsion first chain A residue 220 atom C next chain A residue 220 atom CA next chain A residue 220 atom C next chain A residue 221 atom N value -60 sigma 10 period 1 regards Garib On 8 Jan 2009, at 18:14, Eckhard Hofmann wrote: Hi Phil, sorry, haven't read you question properly. No idea how to get easily from top/par to cif for refmac. Probably would need a little scripting, but that's been exactly your question ... Eckhard XPLO2D from the USF-Suite does this: snippet from manual You feed it a PDB file of the model to which you want to restrain your refinement model (e.g., that high-resolution native structure you already have, even though it may be in a different spacegroup or with different domain orientations). The program generates an X-PLOR include file which contains DIHEdral statements for the PHI, PSI, CHI-1 and CHI-2 torsions of the protein. If you protein contains a hinge region, simply remove or comment-out the relevant PHI and PSI restraints. If you don't want to impose restraints on CHI-1 and/or CHI-2, set the corresponding weights to zero (at the top of the X-PLOR include file). I never tried, but this will be compatible with phenix as well. Cheers Eckhard Phil Evans schrieb: Does anyone have a good way of imposing secondary structure restraints in a low resolution refinement? I've done this in the past as hydrogen bond distance restraints within helices, input to refmac as LINKs , with the list generated with a little program and certain amount of pain refmac now accepts an explicit list of external restraints, as does phenix.refine, but I'm looking for a way of generating these lists for quite a large structure without too much hackery, perhaps from a hydrogen-bond or secondary structure assignment program. Helices are reasonably straightforward (I can see how to do them from eg DSSP), but sheets are more complicated. Any suggestions? I'm sure that someone must have done this Phil -- Eckhard Hofmann eckhard.hofm...@bph.ruhr-uni-bochum.de Ruhr-Uni Bochum AG Proteinkristallographie, LS Biophysik, ND04/316 44780 Bochum Tel: +49-(0)234/32-24463, Sekr. -24461, FAX: -14762
Re: [ccp4bb] Secondary structure restraints
Phil, I have a student who has been working on a python script that will allow the user to manually define hydrogen bonds in pymol (i.e. click on the nitrogen and oxygen atoms that you want to restrain). It then outputs a restraints definition file for refinement in phenix. It can be tedious to define the restraints for a large structure (we are working on a low resolution refinement of a large structure right now), but I haven't been able to find a better alternative. If you would like to play with our scripts, let me know. Sean Johnson Phil Evans wrote: Does anyone have a good way of imposing secondary structure restraints in a low resolution refinement? I've done this in the past as hydrogen bond distance restraints within helices, input to refmac as LINKs , with the list generated with a little program and certain amount of pain refmac now accepts an explicit list of external restraints, as does phenix.refine, but I'm looking for a way of generating these lists for quite a large structure without too much hackery, perhaps from a hydrogen-bond or secondary structure assignment program. Helices are reasonably straightforward (I can see how to do them from eg DSSP), but sheets are more complicated. Any suggestions? I'm sure that someone must have done this Phil -- Sean Johnson, PhD R. Gaurth Hansen Assistant Professor Utah State University Department of Chemistry and Biochemistry 0300 Old Main Hill Logan, UT 84322-0300 (435) 797-2089 (435) 797-3390 (fax) sean.john...@usu.edu
Re: [ccp4bb] Secondary structure restraints
I put together a simple perl script to take WHATIF optimal hydrogen bonds from a known structure and generate refmac or cns restraints. You can limit it to backbone or all h-bonds. Refmac: http://hood.icmb.utexas.edu/~paul/ccp4_hbond CNS: http://hood.icmb.utexas.edu/~paul/cns_hbond Phil Evans wrote: Does anyone have a good way of imposing secondary structure restraints in a low resolution refinement? I've done this in the past as hydrogen bond distance restraints within helices, input to refmac as LINKs , with the list generated with a little program and certain amount of pain refmac now accepts an explicit list of external restraints, as does phenix.refine, but I'm looking for a way of generating these lists for quite a large structure without too much hackery, perhaps from a hydrogen-bond or secondary structure assignment program. Helices are reasonably straightforward (I can see how to do them from eg DSSP), but sheets are more complicated. Any suggestions? I'm sure that someone must have done this Phil -- Paul Paukstelis, Ph.D. Research Associate Institute for Cellular and Molecular Biology The University of Texas at Austin P: 512-471-4778, F: 512-232-3420 p...@icmb.utexas.edu
[ccp4bb] secondary structure restraints
I am trying to build and refine a model into 3.6 angstrom Se-met phased maps. What is the best way to define secondary structure restraints for refinement? (hydrogen bonds? backbone torsion angles?) Are there any tools available to help me define restraints for a specified region, or do I have to define each restraint one at a time (which strikes me as a very tedious exercise). Any other thoughts on best practices for low-resolution model building and refinement would also be appreciated. Thanks, Sean -- Sean Johnson, PhD R. Gaurth Hansen Assistant Professor Utah State University Department of Chemistry and Biochemistry 0300 Old Main Hill Logan, UT 84322-0300 (435) 797-2089 (435) 797-3390 (fax) [EMAIL PROTECTED]
Re: [ccp4bb] secondary structure restraints
Hi Sean, Not an answer to your question but have you looked into model building with TEXTAL? I am not sure about 3.6Ang resolution data but it might be worth looking into. Raji -Included Message-- Date: 13-feb-2008 16:54:33 -0500 From: Sean Johnson [EMAIL PROTECTED] To: CCP4BB@JISCMAIL.AC.UK Subject: [ccp4bb] secondary structure restraints I am trying to build and refine a model into 3.6 angstrom Se-met phased maps. What is the best way to define secondary structure restraints for refinement? (hydrogen bonds? backbone torsion angles?) Are there any tools available to help me define restraints for a specified region, or do I have to define each restraint one at a time (which strikes me as a very tedious exercise). Any other thoughts on best practices for low-resolution model building and refinement would also be appreciated. Thanks, Sean -- Sean Johnson, PhD R. Gaurth Hansen Assistant Professor Utah State University Department of Chemistry and Biochemistry 0300 Old Main Hill Logan, UT 84322-0300 (435) 797-2089 (435) 797-3390 (fax) [EMAIL PROTECTED] -End of Included Message--
Re: [ccp4bb] secondary structure restraints
For a (very) low resolution RNA/protein complex in which we had high(er) resolution structures for both components, I used the optimal hydrogen bonds from these structures (WHATIF output) as restraints. I made a couple perl scripts to take this output and generated either CNS or REFMAC restraints. http://hood.icmb.utexas.edu/~paul bottom of the page. Best, --paul Sean Johnson wrote: I am trying to build and refine a model into 3.6 angstrom Se-met phased maps. What is the best way to define secondary structure restraints for refinement? (hydrogen bonds? backbone torsion angles?) Are there any tools available to help me define restraints for a specified region, or do I have to define each restraint one at a time (which strikes me as a very tedious exercise). Any other thoughts on best practices for low-resolution model building and refinement would also be appreciated. Thanks, Sean
