[ccp4bb] Diamond MX BAG Training - April 2012
To all Diamond MX BAG Users, Diamond Light Source will be holding the next training day for MX BAG Users on Wednesday 25th April 2012. The aim of the day is to provide BAG users with sufficient training to be able to operate any of the Diamond MX beamlines efficiently and get the most benefit from their beamtime. It is essential that each BAG sends at least one representative per calendar year. Sessions include: -Data Acquisition & Analysis -Remote Access -Sample Humidity Control (HC1) -Microbeam Crystallography -In Situ Diffraction Registration is free-of-charge with lunch provided on the 25th and accommodation and dinner for the night of the 24th . Travelling expenses within the UK will also be provided. The training is targeted at all BAG members and is not limited to students and post docs. Individuals wishing to register should reply to this e-mail with their name, BAG, contact details, accommodation and any other requirements. Early registration is recommended as places are limited to twenty. Best wishes, James. Dr James Nicholson, Senior Beamline Scientist MX, Diamond Light Source Ltd. Harwell Science and Innovation Campus, Didcot, Oxfordshire, OX11 0DE, UK.
[ccp4bb] B average lower than Wilson
Dear all, I hope you can help me! I solved a structure at 3.0 A res and everything is quite satisfactory, my only concern is the discrepancy between the B factor from Wilson plot (58.8 A^2) and the average B (36.9). It seems strange, usually the opposite occurs... What does it mean? Which may be the reason? What can I check? Some details: Rmerge: 15.0 (51.8) I/σ(I): 7.30 (1.84) 5 monomers/AU R/Rfree: 19.7/23.3 ...ask for other infoes if you need. Any advice/suggestion is welcome!!
Re: [ccp4bb] B average lower than Wilson
-BEGIN PGP SIGNED MESSAGE- Hash: SHA1 Dear , the Wilson plot is only meaningful with data better than 4A. Since your data extends to only 3A (and according to your I/sig(I) stats you pushed that limit a little...), you may have too little data for a good fit. That's only one possibility. Your data may have ice rings resulting in poor intensity estimates, are the atoms may indeed have low B-values and the reason for limiting the resolution may have a different source than usual,... Tim On 03/05/2012 04:16 PM, anna anna wrote: > Dear all, I hope you can help me! > I solved a structure at 3.0 A res and everything is quite satisfactory, my > only concern is the discrepancy between > the B factor from Wilson plot (58.8 A^2) and the average B (36.9). It seems > strange, usually the opposite occurs... > What does it mean? Which may be the reason? What can I check? > Some details: > Rmerge: 15.0 (51.8) > I/σ(I): 7.30 (1.84) > 5 monomers/AU > R/Rfree: 19.7/23.3 > ...ask for other infoes if you need. > > Any advice/suggestion is welcome!! > - -- - -- Dr Tim Gruene Institut fuer anorganische Chemie Tammannstr. 4 D-37077 Goettingen GPG Key ID = A46BEE1A -BEGIN PGP SIGNATURE- Version: GnuPG v1.4.10 (GNU/Linux) Comment: Using GnuPG with Mozilla - http://enigmail.mozdev.org/ iD8DBQFPVOHcUxlJ7aRr7hoRAtnlAKDgx9rk4MZ2qxF2vJRMCJlQhTGlZgCgpQwm 2eOrSTf5N8+a+KLZcV74zks= =z2bK -END PGP SIGNATURE-
Re: [ccp4bb] B average lower than Wilson
Hi, Please see a similar thread at ccp4bb earlier at the following link http://www.proteincrystallography.org/ccp4bb/message16011.html Padayatti 2012/3/5 anna anna : > Dear all, I hope you can help me! > I solved a structure at 3.0 A res and everything is quite satisfactory, my > only concern is the discrepancy between > the B factor from Wilson plot (58.8 A^2) and the average B (36.9). It seems > strange, usually the opposite occurs... > What does it mean? Which may be the reason? What can I check? > Some details: > Rmerge: 15.0 (51.8) > I/σ(I): 7.30 (1.84) > 5 monomers/AU > R/Rfree: 19.7/23.3 > ...ask for other infoes if you need. > > Any advice/suggestion is welcome!! -- Pius S Padayatti,PhD, Phone: 216-658-4528
Re: [ccp4bb] B average lower than Wilson
Dear all, thanks for help! I'm using Refmac without TLS. Tim's explanation is pretty convincing to me. My data have no ice rings compromising the extimation of I, but the map, is more defined then I expect at 3 A resolution, so a "low" average B seems consistent. Which can be other causes for limited resolution? Anna Il giorno 05 marzo 2012 16:56, H. Raaijmakers ha scritto: > Anna, > > Did you use TLS refinement, and are looking at the residual B? > then use tlsanl to add the tls contribution. > > cheers, > > Hans > > anna anna schreef: > > Dear all, I hope you can help me! > > I solved a structure at 3.0 A res and everything is quite satisfactory, > my > > only concern is the discrepancy between > > the B factor from Wilson plot (58.8 A^2) and the average B (36.9). It > > seems > > strange, usually the opposite occurs... > > What does it mean? Which may be the reason? What can I check? > > Some details: > > Rmerge: 15.0 (51.8) > > I/ó(I): 7.30 (1.84) > > 5 monomers/AU > > R/Rfree: 19.7/23.3 > > ...ask for other infoes if you need. > > > > Any advice/suggestion is welcome!! > > > > >
Re: [ccp4bb] sudden drop in R/Rfree
Dear All, Thanks for all the suggestion. Data is anisotropic. Since anisotropic correction with the UCLA server didnt give any good Molprobity scores and didnt reduce the R/Rfree at 2.1A, I am using ~3.3A data for further refinement. Using Tom's suggestion of target restrains reduced the gap brtween R/Rfee to with in 5%. I haven't tried Pavel' suggestion but when I do I will post the results. I used a 2.3 A MR model as target to get R/Rfree 0.2209/0.2597. When I used coot real space refine zone to fix few molprobity outliers, most of the favored residues become outliers. Almost all residues changed to become outliers. I am wondering why all the residues are moving from favored to outlier regions after restrained refinement. I thought all the residues would be fixed (fit) to the map and wouldn't change during the coot exercise, and this is what I see in the 2.3 A target/model structure ( with R/Rfree 0.1693/0.1961). I don't know if is this common for a low resolution or I didn't understand this whole thing of refinement correctly. Thanks for all the help, Regards,Raj > Date: Sun, 4 Mar 2012 21:02:51 + > From: eleanor.dod...@york.ac.uk > Subject: Re: [ccp4bb] sudden drop in R/Rfree > To: CCP4BB@JISCMAIL.AC.UK > > Presumably your data is quite anisotropic, and low resolution, so it is > quite likely that a TLS model will give much better description of the B > factors than more classical refinement. > > Modelling solvent at that resolotion will be tricky of course. > Elesnor > > > > On Mar 4 2012, Joseph Cockburn wrote: > > >Hi Rajesh, > >If you're seeing a lot of extra density coming up in the map in regions > >where you previously added waters, is it possible that this extra density > >corresponds to a part of your protein that you previously thought was > >disordered and is thus missing from the current model? At this resolution > >you wouldn't expect to see many waters. > >Also, to the best of my knowledge, the relative weighting of the X-ray and > >geometry terms in BUSTER is set by the program so as to produce a rmsd in > >bond lengths equal to a target value. The default value of this is 0.01 > >Ang (I think) but you can change this using the -r option on the command > >line. Using a lower value will reduce the weight on the X-ray term and may > >lower the R/R-free gap. > >Best wishes, > >Joe > > > > > > > >> > >> > >> Dear All, I have a 3.3 A data for a protein whose SG is P6522. Model > >> used was wild type structure of same protein at 2.3 A. After molecular > >> replacement, first three rounds of refinement the R/Rf was 26/32.8, > >> 27.1/31.72 % and 7.35/30.88 % respectively.In the fourth round I refined > >> with TLS and NCS abd added water and the R/Rf dropped to 19.34/26.46. It > >> has almost 7% difference. I also see lot of unanswerable density in the > >> map where lot of waters were placed. Model fits to the map like a low > >> resolution data with most of side chains don't have best density. I was > >> not expecting such a sudden drop in the R/Rfree and a difference is > >> 7.2%. I am wondering if I am in right direction. I am not sure if this > >> usual for 3.3A data or in general any data if we consider the > >> difference. I appreciate your valuable suggestions. ThanksRaj > >> > >> > > > > -- > Professor Eleanor Dodson > YSNL, Dept of Chemistry > University of York > Heslington YO10 5YW > tel: 00 44 1904 328259 > Fax: 00 44 1904 328266
[ccp4bb] REFMAC Riding Hydrogens
Hello CCP4 community, I'm posting at large regarding a previously raised issue for REFMAC for which I cannot find the conclusion in the old threads. Specifically, does REFMAC add riding hydrogens during default refinement? Though I've not requested that any hydrogens be added, only used if in the input, I see the following in the output header: REMARK 3 HYDROGENS HAVE BEEN ADDED IN THE RIDING POSITIONS Does anyone know exactly what this remark is referring to? Are riding hydrogens added for stereochemical restraints, or are their scattering contributions calculated regardless of they are explicitly defined? Can the hydrogen behavior in REFMAC be more explicitly controlled. Thanks, --Paul
Re: [ccp4bb] REFMAC Riding Hydrogens
Hi Paul - Refmac does indeed add riding hydrogens by default; see here: http://www.ccp4.ac.uk/dist/html/refmac5/keywords/restraints.html#makecif The REMARK 3 is referring to what you think it is: riding hydrogens have been added. (They are not, however, written to the output file unless you request that.) I believe the H atoms are used for both stereochemistry and scattering, although there may be some fudging on the scattering factors. Adding the riding hydrogens generally gives you some improvement in R factors even with a good quality (i.e. stereochemically correct) model. If you look at the link I gave above, you will see that you can do three things with hydrogens: never use them, use the riding hydrogens, or only use H atoms given in the input file (which you could of course tailor to your needs). Hope that helps, Matt On 3/5/12 2:15 PM, Paul Smith wrote: Hello CCP4 community, I'm posting at large regarding a previously raised issue for REFMAC for which I cannot find the conclusion in the old threads. Specifically, does REFMAC add riding hydrogens during default refinement? Though I've not requested that any hydrogens be added, only used if in the input, I see the following in the output header: REMARK 3 HYDROGENS HAVE BEEN ADDED IN THE RIDING POSITIONS Does anyone know exactly what this remark is referring to? Are riding hydrogens added for stereochemical restraints, or are their scattering contributions calculated regardless of they are explicitly defined? Can the hydrogen behavior in REFMAC be more explicitly controlled. Thanks, --Paul -- Matthew Franklin, Ph. D. Senior Research Scientist New York Structural Biology Center 89 Convent Avenue, New York, NY 10027 (646) 275-7165
Re: [ccp4bb] REFMAC Riding Hydrogens
On the interface under Refinement Parameters there are three options 1) Use if present 2) Do not use 3) Generate all if you are using script then default is generate all (add them riding positions). You can switch to other options: make hydr all# generate all - default option make hydr yes # use if present in the file make hydr no # do not use Controlling output make hout yes # write hydrogens to the output file make hout no # do not write - default optoon I hope it helps regards Garib On 5 Mar 2012, at 19:15, Paul Smith wrote: > Hello CCP4 community, > > I'm posting at large regarding a previously raised issue for REFMAC for which > I cannot find the conclusion in the old threads. > > Specifically, does REFMAC add riding hydrogens during default refinement? > > Though I've not requested that any hydrogens be added, only used if in the > input, I see the following in the output header: > > REMARK 3 HYDROGENS HAVE BEEN ADDED IN THE RIDING POSITIONS > > Does anyone know exactly what this remark is referring to? > > Are riding hydrogens added for stereochemical restraints, or are their > scattering contributions calculated regardless of they are explicitly defined? > > Can the hydrogen behavior in REFMAC be more explicitly controlled. > > Thanks, > > --Paul > > Garib N Murshudov Structural Studies Division MRC Laboratory of Molecular Biology Hills Road Cambridge CB2 0QH UK Email: ga...@mrc-lmb.cam.ac.uk Web http://www.mrc-lmb.cam.ac.uk
Re: [ccp4bb] REFMAC Riding Hydrogens
Hi, On Mon, Mar 5, 2012 at 11:52 AM, Matthew Franklin wrote: > Adding the riding hydrogens generally gives you some improvement in R > factors even with a good quality (i.e. stereochemically correct) model. > and here are the results of more or less systematic test that prove this: see "On contribution of hydrogen atoms to X-ray scattering" here: http://www.phenix-online.org/newsletter/ Pavel
Re: [ccp4bb] REFMAC Riding Hydrogens
-BEGIN PGP SIGNED MESSAGE- Hash: SHA1 Dear Pavel, you may want to add to the structures mentioned in [1] one or two organic structures present in the Cambridge Database. "Until recently it was customary to ignore hydrogen atoms throughout the process of crystallographic X-‐ray structure determination." [1] 'recently' as in 1997 [2]? Even though 1997 is probably a poor estimation of the corresponding year... Cheers, Tim [1] "On contribution of hydrogen atoms to X-ray scattering" http://www.phenix-online.org/newsletter/ [2] http://shelx.uni-ac.gwdg.de/SHELX/shelx.pdf On 03/05/2012 09:14 PM, Pavel Afonine wrote: > Hi, > > On Mon, Mar 5, 2012 at 11:52 AM, Matthew Franklin wrote: > >> Adding the riding hydrogens generally gives you some improvement in R >> factors even with a good quality (i.e. stereochemically correct) model. >> > > and here are the results of more or less systematic test that prove this: > > see "On contribution of hydrogen atoms to X-ray scattering" > here: > http://www.phenix-online.org/newsletter/ > > Pavel > - -- Dr Tim Gruene Institut fuer anorganische Chemie Tammannstr. 4 D-37077 Goettingen GPG Key ID = A46BEE1A -BEGIN PGP SIGNATURE- Version: GnuPG v1.4.11 (GNU/Linux) Comment: Using GnuPG with Mozilla - http://enigmail.mozdev.org/ iD8DBQFPVSXkUxlJ7aRr7hoRAm1TAJ9Hyfhkl3yhD5QSKw9I4RSK58m0fACgmlxk YGILzeMam/3gQVmCeh0vQ8k= =3m2J -END PGP SIGNATURE-
Re: [ccp4bb] REFMAC Riding Hydrogens
Dear Tim, good catch, thanks; I could craft that phrase more carefully! Although often it may not be quite fair to take phrases out of context: this newsletter article was written in the context of macromolecular refinement. And yes, "recently" may be a broad term -:) All the best, Pavel On Mon, Mar 5, 2012 at 12:45 PM, Tim Gruene wrote: > -BEGIN PGP SIGNED MESSAGE- > Hash: SHA1 > > Dear Pavel, > > you may want to add to the structures mentioned in [1] one or two > organic structures present in the Cambridge Database. > > "Until recently it was customary to ignore hydrogen atoms throughout the > process of crystallographic X-‐ray structure determination." [1] > > 'recently' as in 1997 [2]? Even though 1997 is probably a poor > estimation of the corresponding year... > > Cheers, > Tim > > > [1] "On contribution of hydrogen atoms to X-ray scattering" > http://www.phenix-online.org/newsletter/ > [2] http://shelx.uni-ac.gwdg.de/SHELX/shelx.pdf > > On 03/05/2012 09:14 PM, Pavel Afonine wrote: > > Hi, > > > > On Mon, Mar 5, 2012 at 11:52 AM, Matthew Franklin >wrote: > > > >> Adding the riding hydrogens generally gives you some improvement in R > >> factors even with a good quality (i.e. stereochemically correct) model. > >> > > > > and here are the results of more or less systematic test that prove this: > > > > see "On contribution of hydrogen atoms to X-ray scattering" > > here: > > http://www.phenix-online.org/newsletter/ > > > > Pavel > > > > - -- > Dr Tim Gruene > Institut fuer anorganische Chemie > Tammannstr. 4 > D-37077 Goettingen > > GPG Key ID = A46BEE1A > -BEGIN PGP SIGNATURE- > Version: GnuPG v1.4.11 (GNU/Linux) > Comment: Using GnuPG with Mozilla - http://enigmail.mozdev.org/ > > iD8DBQFPVSXkUxlJ7aRr7hoRAm1TAJ9Hyfhkl3yhD5QSKw9I4RSK58m0fACgmlxk > YGILzeMam/3gQVmCeh0vQ8k= > =3m2J > -END PGP SIGNATURE- >
Re: [ccp4bb] B average lower than Wilson
Not an answer, but if you look at the Wilson plot after scala or truncate, you can see how good the estimate is - at 3A those can be pretty awful Eleanot On Mar 5 2012, anna anna wrote: Dear all, I hope you can help me! I solved a structure at 3.0 A res and everything is quite satisfactory, my only concern is the discrepancy between the B factor from Wilson plot (58.8 A^2) and the average B (36.9). It seems strange, usually the opposite occurs... What does it mean? Which may be the reason? What can I check? Some details: Rmerge: 15.0 (51.8) I/σ(I): 7.30 (1.84) 5 monomers/AU R/Rfree: 19.7/23.3 ...ask for other infoes if you need. Any advice/suggestion is welcome!! -- Professor Eleanor Dodson YSNL, Dept of Chemistry University of York Heslington YO10 5YW tel: 00 44 1904 328259 Fax: 00 44 1904 328266
Re: [ccp4bb] sudden drop in R/Rfree
No comments on much of your message but I cannot believe you can get better results by cutting anisotropic 2.1A data to 3.3A!! You MUST be throwing away valuable experimental information,, remember crystallography is not about getting low R factors but about getting a model consistent with your experiment. eleanor On Mar 5 2012, Rajesh kumar wrote: Dear All, Thanks for all the suggestion. Data is anisotropic. Since anisotropic correction with the UCLA server didnt give any good Molprobity scores and didnt reduce the R/Rfree at 2.1A, I am using ~3.3A data for further refinement. Using Tom's suggestion of target restrains reduced the gap brtween R/Rfee to with in 5%. I haven't tried Pavel' suggestion but when I do I will post the results. I used a 2.3 A MR model as target to get R/Rfree 0.2209/0.2597. When I used coot real space refine zone to fix few molprobity outliers, most of the favored residues become outliers. Almost all residues changed to become outliers. I am wondering why all the residues are moving from favored to outlier regions after restrained refinement. I thought all the residues would be fixed (fit) to the map and wouldn't change during the coot exercise, and this is what I see in the 2.3 A target/model structure ( with R/Rfree 0.1693/0.1961). I don't know if is this common for a low resolution or I didn't understand this whole thing of refinement correctly. Thanks for all the help, Regards,Raj Date: Sun, 4 Mar 2012 21:02:51 + From: eleanor.dod...@york.ac.uk Subject: Re: [ccp4bb] sudden drop in R/Rfree To: CCP4BB@JISCMAIL.AC.UK Presumably your data is quite anisotropic, and low resolution, so it is quite likely that a TLS model will give much better description of the B factors than more classical refinement. Modelling solvent at that resolotion will be tricky of course. Elesnor On Mar 4 2012, Joseph Cockburn wrote: > Hi Rajesh, If you're seeing a lot of extra density coming up in the > map in regions where you previously added waters, is it possible that > this extra density corresponds to a part of your protein that you > previously thought was disordered and is thus missing from the current > model? At this resolution you wouldn't expect to see many waters. > Also, to the best of my knowledge, the relative weighting of the X-ray > and geometry terms in BUSTER is set by the program so as to produce a > rmsd in bond lengths equal to a target value. The default value of > this is 0.01 Ang (I think) but you can change this using the -r option > on the command line. Using a lower value will reduce the weight on the > X-ray term and may lower the R/R-free gap. Best wishes, Joe > > > >> >> >> Dear All, I have a 3.3 A data for a protein whose SG is P6522. Model >> used was wild type structure of same protein at 2.3 A. After >> molecular replacement, first three rounds of refinement the R/Rf was >> 26/32.8, 27.1/31.72 % and 7.35/30.88 % respectively.In the fourth >> round I refined with TLS and NCS abd added water and the R/Rf dropped >> to 19.34/26.46. It has almost 7% difference. I also see lot of >> unanswerable density in the map where lot of waters were placed. >> Model fits to the map like a low resolution data with most of side >> chains don't have best density. I was not expecting such a sudden >> drop in the R/Rfree and a difference is 7.2%. I am wondering if I am >> in right direction. I am not sure if this usual for 3.3A data or in >> general any data if we consider the difference. I appreciate your >> valuable suggestions. ThanksRaj >> >> > -- Professor Eleanor Dodson YSNL, Dept of Chemistry University of York Heslington YO10 5YW tel: 00 44 1904 328259 Fax: 00 44 1904 328266 -- Professor Eleanor Dodson YSNL, Dept of Chemistry University of York Heslington YO10 5YW tel: 00 44 1904 328259 Fax: 00 44 1904 328266
