Re: [ccp4bb] Charge flipping.
Dear Francis, I am very interested in your work with this. In the paper :- A. Mukherjee, J.R. Helliwell and P. Main 'The use of MULTAN to locate the positions of anomalous scatterers'. (1989) Acta Cryst. *A45*, 715-718. you will see that missing centric reflections was not limiting. However, the more dilute the anomalous scatterers versus the protein atoms led to the practical consequence of increasing the number of required iterations to a successful answer. That said the potential effect of missing centric data and the influence of random and systematic errors on the charge flipping may of course be different. Best wishes, John Prof John R helliwell On Sat, May 22, 2010 at 3:23 PM, Francis E Reyes wrote: > Hi all > > I've been playing around with charge flipping for macromolecular > substructure determination with pretty promising results. I'm particularly > attracted to the fact that it solves structures in P1, with no space group > assumptions and curious how it would handle some of the pseudosymmetry cases > I've come into in my time. > > I'd like to know if anyone's had experience with this method, and open up > the discussion with the following questions: > > As the algorithm starts with completely random phases and charge flips the > map in P1, what is the importance of measuring (good or any) anomalous > signal at all (for the sole purpose of finding the heavy atoms)? At first > pass it would seem that just as long as you have an incorporated heavy atom > and the density of that region is greater than delta, that this alone would > be sufficient for locating the position of the heavy atom. In other words > just as long as your heavy atom is sufficiently higher in contrast than your > protein/rna it would be a good enough criteria. > > In the above regime, would the importance of measuring anomalous data be > more important for substructure refinement (via phaser, mlphare, sharp, > solve/resolve)? > > Now to a more specific question for those who've had experience (or maybe > the authors are subscribed here): > > Orthorhombic C2221 using SUPERFLIP heavy atoms are found with great > peakiness (before noise suppression: peakiness = 5, after noise suppression > peakiness >25, good separation of heavy atom peaks from noise peaks in > resulting pdb). Yet the space group check via the sym operators is rather > poor (overall agreement close to 100). My interpretation is that the heavy > atoms are found, but the space group is wrong? > > > > Thanks! > > F > > - > Francis Reyes M.Sc. > 215 UCB > University of Colorado at Boulder > > gpg --keyserver pgp.mit.edu --recv-keys 67BA8D5D > > 8AE2 F2F4 90F7 9640 28BC 686F 78FD 6669 67BA 8D5D > -- Professor John R Helliwell DSc
Re: [ccp4bb] Charge flipping.
Dear all, We have tested the Charge Flipping algorithm with SUPERFLIP program on various experimental data (anomalous delta-F's, MAD FA's). see http://www.cbs.cnrs.fr/SP/crystal/SUPERFLIP/ & Dumas, van der Lee, Acta Cryst D64, 864-73 In all successfull trials, a good quality substructure is obtained. As outlined below, the main requirements are good completion and high quality experimental data! This is a relatively new approach and there's still room for improvement. > Francis E Reyes wrote: > Orthorhombic C2221 using SUPERFLIP heavy atoms are found, ... > but the SG is wrong. Yes, this is the most probable diagnostic hypothesis. It looks like you have missed a space group. This ambiguity can arise with twinning, NCS in special position,... I had a similar case for Se substrcture solution using SUPERFLIP with data indexed as C2221, where P21 lattice approximates C2221. SUPERFLIP (http://superflip.fzu.cz/) can try to derive the symmetry operations of the P1 charge flipping density, independently of the symmetry entered in the input file (see Palatinus, J; applied cryst, 41,975-84) Restart a new job using the keyword derivesymmetry: the list of symmetry operations, centering vectors will be displayed (with agreement factors) in the .sflog file -'derivesymmetry no' the output density is in P1, no shifting or averaging or -'derivesymmetry use' shifting and averaging the output density. If you send me the inputs and sflog files I will have a closer look. Christian. -- Christian Dumas, Centre de Biochimie Structurale, 34090 Montpellier Cedex, France Tel: +33-(0)467.417.705 George M. Sheldrick wrote: > I have also played with charge flipping and my experience was the same as > Kevin's. Michael Woolfson once said that all 'direct' methods work fine with > perfect data. The method requires expansion of the data to P1 which seems > to degrade the quality of the solution; when the data are noisy, as is > usually the case with real anomalous differences, the space group can be a > useful constraint, as is the number of sites expected. If necessary one > can easily run the usual programs in all potential space groups. For the > solution of small molecule structures using atomic resolution (say 0.9A) > native data, the data are much less borderline and charge flipping in P1 > is a good way to explore phase space. > Another reason why charge flipping may not work so well with real > anomalous differences is that the data tend to be rather incomplete, > for example all the centric reflections are missing. This degrades > the quality of the resulting maps, which is more serious if you are > modifying low densities than when you are just searching for peaks. George Prof. George M. Sheldrick FRS Dept. Structural Chemistry, University of Goettingen, Tammannstr. 4, D37077 Goettingen, Germany Tel. +49-551-39-3021 or -3068 Fax. +49-551-39-22582 Kewin Cowtan wrote: > I played with this (coded from scratch, both simple algorithm and a few > tweaks) for a couple of weeks for solving heavy atom substructures. With > perfect FAs it works well and quickly. With real delta-F's it didn't > work at all. Can't remember if I tried perfect delta-F's. > Probably SUPERFLIP is better than my quick implementation though. Francis E Reyes wrote: > Hi all > I've been playing around with charge flipping for macromolecular > substructure determination with pretty promising results. I'm > particularly attracted to the fact that it solves structures in P1, > with no space group assumptions and curious how it would handle some > of the pseudosymmetry cases I've come into in my time. > > I'd like to know if anyone's had experience with this method, and > open up the discussion with the following questions: > > As the algorithm starts with completely random phases and charge > flips the map in P1, what is the importance of measuring (good or > any) anomalous signal at all (for the sole purpose of finding the > heavy atoms)? At first pass it would seem that just as long as you > have an incorporated heavy atom and the density of that region is > greater than delta, that this alone would be sufficient for locating > the position of the heavy atom. In other words just as long as your > heavy atom is sufficiently higher in contrast than your protein/rna it > would be a good enough criteria. > > In the above regime, would the importance of measuring anomalous data > be more important for substructure refinement (via phaser, mlphare, > sharp, solve/resolve)? > > Now to a more specific question for those who've had experience (or > maybe the authors are subscribed here): > > Orthorhombic C2221 using SUPERFLIP heavy atoms are found with great > peakiness (before noise suppression: peakiness = 5, after noise > suppression peakiness >25, good separation of heavy atom peaks from > noise peaks in resulting pdb). Yet the space group check via the sym > operators is rather poor (overall agreement close to 100).
Re: [ccp4bb] Charge flipping - PS.
Another reason why charge flipping may not work so well with real anomalous differences is that the data tend to be rather incomplete, for example all the centric reflections are missing. This degrades the quality of the resulting maps, which is more serious if you are modifying low densities than when you are just searching for peaks. George Prof. George M. Sheldrick FRS Dept. Structural Chemistry, University of Goettingen, Tammannstr. 4, D37077 Goettingen, Germany Tel. +49-551-39-3021 or -3068 Fax. +49-551-39-22582 On Mon, 24 May 2010, George M. Sheldrick wrote: > I have also played with charge flipping and my experience was the same as > Kevin's. Michael Woolfson once said that all 'direct' methods work fine with > perfect data. The method requires expansion of the data to P1 which seems > to degrade the quality of the solution; when the data are noisy, as is > usually the case with real anomalous differences, the space group can be a > useful constraint, as is the number of sites expected. If necessary one > can easily run the usual programs in all potential space groups. For the > solution of small molecule structures using atomic resolution (say 0.9A) > native data, the data are much less borderline and charge flipping in P1 > is a good way to explore phase space. > > George > > Prof. George M. Sheldrick FRS > Dept. Structural Chemistry, > University of Goettingen, > Tammannstr. 4, > D37077 Goettingen, Germany > Tel. +49-551-39-3021 or -3068 > Fax. +49-551-39-22582 > > > On Mon, 24 May 2010, Kevin Cowtan wrote: > > > I played with this (coded from scratch, both simple algorithm and a few > > tweaks) for a couple of weeks for solving heavy atom substructures. With > > perfect FAs it works well and quickly. With real delta-F's it didn't work at > > all. Can't remember if I tried perfect delta-F's. > > > > Probably SUPERFLIP is better than my quick implementation though. > > > > Francis E Reyes wrote: > > > Hi all > > > > > > I've been playing around with charge flipping for macromolecular > > > substructure determination with pretty promising results. I'm particularly > > > attracted to the fact that it solves structures in P1, with no space group > > > assumptions and curious how it would handle some of the pseudosymmetry > > > cases > > > I've come into in my time. > > > > > > I'd like to know if anyone's had experience with this method, and open up > > > the discussion with the following questions: > > > > > > As the algorithm starts with completely random phases and charge flips > > > the > > > map in P1, what is the importance of measuring (good or any) anomalous > > > signal at all (for the sole purpose of finding the heavy atoms)? At first > > > pass it would seem that just as long as you have an incorporated heavy > > > atom > > > and the density of that region is greater than delta, that this alone > > > would > > > be sufficient for locating the position of the heavy atom. In other words > > > just as long as your heavy atom is sufficiently higher in contrast than > > > your > > > protein/rna it would be a good enough criteria. > > > > > > In the above regime, would the importance of measuring anomalous data be > > > more important for substructure refinement (via phaser, mlphare, sharp, > > > solve/resolve)? > > > > > > Now to a more specific question for those who've had experience (or maybe > > > the authors are subscribed here): > > > > > > Orthorhombic C2221 using SUPERFLIP heavy atoms are found with great > > > peakiness (before noise suppression: peakiness = 5, after noise > > > suppression > > > peakiness >25, good separation of heavy atom peaks from noise peaks in > > > resulting pdb). Yet the space group check via the sym operators is rather > > > poor (overall agreement close to 100). My interpretation is that the > > > heavy > > > atoms are found, but the space group is wrong? > > > > > > > > > > > > Thanks! > > > > > > F > > > > > > - > > > Francis Reyes M.Sc. > > > 215 UCB > > > University of Colorado at Boulder > > > > > > gpg --keyserver pgp.mit.edu --recv-keys 67BA8D5D > > > > > > 8AE2 F2F4 90F7 9640 28BC 686F 78FD 6669 67BA 8D5D > > > > > >
Re: [ccp4bb] Charge flipping.
I have also played with charge flipping and my experience was the same as Kevin's. Michael Woolfson once said that all 'direct' methods work fine with perfect data. The method requires expansion of the data to P1 which seems to degrade the quality of the solution; when the data are noisy, as is usually the case with real anomalous differences, the space group can be a useful constraint, as is the number of sites expected. If necessary one can easily run the usual programs in all potential space groups. For the solution of small molecule structures using atomic resolution (say 0.9A) native data, the data are much less borderline and charge flipping in P1 is a good way to explore phase space. George Prof. George M. Sheldrick FRS Dept. Structural Chemistry, University of Goettingen, Tammannstr. 4, D37077 Goettingen, Germany Tel. +49-551-39-3021 or -3068 Fax. +49-551-39-22582 On Mon, 24 May 2010, Kevin Cowtan wrote: > I played with this (coded from scratch, both simple algorithm and a few > tweaks) for a couple of weeks for solving heavy atom substructures. With > perfect FAs it works well and quickly. With real delta-F's it didn't work at > all. Can't remember if I tried perfect delta-F's. > > Probably SUPERFLIP is better than my quick implementation though. > > Francis E Reyes wrote: > > Hi all > > > > I've been playing around with charge flipping for macromolecular > > substructure determination with pretty promising results. I'm particularly > > attracted to the fact that it solves structures in P1, with no space group > > assumptions and curious how it would handle some of the pseudosymmetry cases > > I've come into in my time. > > > > I'd like to know if anyone's had experience with this method, and open up > > the discussion with the following questions: > > > > As the algorithm starts with completely random phases and charge flips the > > map in P1, what is the importance of measuring (good or any) anomalous > > signal at all (for the sole purpose of finding the heavy atoms)? At first > > pass it would seem that just as long as you have an incorporated heavy atom > > and the density of that region is greater than delta, that this alone would > > be sufficient for locating the position of the heavy atom. In other words > > just as long as your heavy atom is sufficiently higher in contrast than your > > protein/rna it would be a good enough criteria. > > > > In the above regime, would the importance of measuring anomalous data be > > more important for substructure refinement (via phaser, mlphare, sharp, > > solve/resolve)? > > > > Now to a more specific question for those who've had experience (or maybe > > the authors are subscribed here): > > > > Orthorhombic C2221 using SUPERFLIP heavy atoms are found with great > > peakiness (before noise suppression: peakiness = 5, after noise suppression > > peakiness >25, good separation of heavy atom peaks from noise peaks in > > resulting pdb). Yet the space group check via the sym operators is rather > > poor (overall agreement close to 100). My interpretation is that the heavy > > atoms are found, but the space group is wrong? > > > > > > > > Thanks! > > > > F > > > > - > > Francis Reyes M.Sc. > > 215 UCB > > University of Colorado at Boulder > > > > gpg --keyserver pgp.mit.edu --recv-keys 67BA8D5D > > > > 8AE2 F2F4 90F7 9640 28BC 686F 78FD 6669 67BA 8D5D > >
Re: [ccp4bb] Charge flipping.
I played with this (coded from scratch, both simple algorithm and a few tweaks) for a couple of weeks for solving heavy atom substructures. With perfect FAs it works well and quickly. With real delta-F's it didn't work at all. Can't remember if I tried perfect delta-F's. Probably SUPERFLIP is better than my quick implementation though. Francis E Reyes wrote: Hi all I've been playing around with charge flipping for macromolecular substructure determination with pretty promising results. I'm particularly attracted to the fact that it solves structures in P1, with no space group assumptions and curious how it would handle some of the pseudosymmetry cases I've come into in my time. I'd like to know if anyone's had experience with this method, and open up the discussion with the following questions: As the algorithm starts with completely random phases and charge flips the map in P1, what is the importance of measuring (good or any) anomalous signal at all (for the sole purpose of finding the heavy atoms)? At first pass it would seem that just as long as you have an incorporated heavy atom and the density of that region is greater than delta, that this alone would be sufficient for locating the position of the heavy atom. In other words just as long as your heavy atom is sufficiently higher in contrast than your protein/rna it would be a good enough criteria. In the above regime, would the importance of measuring anomalous data be more important for substructure refinement (via phaser, mlphare, sharp, solve/resolve)? Now to a more specific question for those who've had experience (or maybe the authors are subscribed here): Orthorhombic C2221 using SUPERFLIP heavy atoms are found with great peakiness (before noise suppression: peakiness = 5, after noise suppression peakiness >25, good separation of heavy atom peaks from noise peaks in resulting pdb). Yet the space group check via the sym operators is rather poor (overall agreement close to 100). My interpretation is that the heavy atoms are found, but the space group is wrong? Thanks! F - Francis Reyes M.Sc. 215 UCB University of Colorado at Boulder gpg --keyserver pgp.mit.edu --recv-keys 67BA8D5D 8AE2 F2F4 90F7 9640 28BC 686F 78FD 6669 67BA 8D5D
[ccp4bb] Charge flipping.
Hi all I've been playing around with charge flipping for macromolecular substructure determination with pretty promising results. I'm particularly attracted to the fact that it solves structures in P1, with no space group assumptions and curious how it would handle some of the pseudosymmetry cases I've come into in my time. I'd like to know if anyone's had experience with this method, and open up the discussion with the following questions: As the algorithm starts with completely random phases and charge flips the map in P1, what is the importance of measuring (good or any) anomalous signal at all (for the sole purpose of finding the heavy atoms)? At first pass it would seem that just as long as you have an incorporated heavy atom and the density of that region is greater than delta, that this alone would be sufficient for locating the position of the heavy atom. In other words just as long as your heavy atom is sufficiently higher in contrast than your protein/rna it would be a good enough criteria. In the above regime, would the importance of measuring anomalous data be more important for substructure refinement (via phaser, mlphare, sharp, solve/resolve)? Now to a more specific question for those who've had experience (or maybe the authors are subscribed here): Orthorhombic C2221 using SUPERFLIP heavy atoms are found with great peakiness (before noise suppression: peakiness = 5, after noise suppression peakiness >25, good separation of heavy atom peaks from noise peaks in resulting pdb). Yet the space group check via the sym operators is rather poor (overall agreement close to 100). My interpretation is that the heavy atoms are found, but the space group is wrong? Thanks! F - Francis Reyes M.Sc. 215 UCB University of Colorado at Boulder gpg --keyserver pgp.mit.edu --recv-keys 67BA8D5D 8AE2 F2F4 90F7 9640 28BC 686F 78FD 6669 67BA 8D5D
