Re: [ccp4bb] High R/Rfree after MR
Dear Randy, On Fri, Oct 13, 2017 at 04:11:44PM +0100, Randy Read wrote: > Just to add to this point. The MR algorithms in Phaser are now able > to make better use of intensity data, which is particularly > important when you have any very weak data. Having weak data can’t > be avoided when you have serious anisotropy (or tNCS or a > combination of the two). Unfortunately, if you use amplitudes that > have been through the French & Wilson (truncate) algorithm, the real > variation in intensity is partially masked because the posterior > amplitude values are computed on the prior assumption that all the > reflections in a resolution shell have the same underlying intensity > distribution. Your "unfortunately" calsue may be the case with the UCLA server, but your statement is not true about what the STARANISO server (or STARANISO as invoked within autoPROC) does: as I already indicated in a reply to you a few months ago in this BB, the version of TRUNCATE in STARANISO applies the French & Wilson procedure with a prior Wilson probability whose expectation value for the intensity is modulated by the anisotropy of the dataset. This is clearly explained on the server - see http://staraniso.globalphasing.org/staraniso_about.html#step16 With best wishes, Gerard. -- > The UCLA server actually uses Phaser under the hood — what they add is to > turn the anisotropic B-values into suggested resolution limits in the > different directions. However, I don’t think they allow you yet to submit > intensities, which would be better. > > Best wishes, > > Randy Read > > - > Randy J. Read > Department of Haematology, University of Cambridge > Cambridge Institute for Medical ResearchTel: +44 1223 336500 > Wellcome Trust/MRC Building Fax: +44 1223 336827 > Hills RoadE-mail: > rj...@cam.ac.uk > Cambridge CB2 0XY, U.K. > www-structmed.cimr.cam.ac.uk > > > On 13 Oct 2017, at 10:29, vincent Chaptal wrote: > > > > Dear Gia and Paul, > > > > about anisotropy, one point to keep in mind is that it is not necessarily > > linked to the difference in resolution limits. > > In fact I am at the moment working on one of these cases, with extremely > > large difference in resolution limits, but relatively low anisotropy. > > Anisotropy is more a deviation from "normal" intensity falloff as a > > function of resolution. There is a thin difference/relationship between the > > two concepts that I think is worth investigating. > > > > we have performed a statistical analysis of this phenomenon and the paper > > is in revision at the moment, but if you want to know where your anisotropy > > stands in respect to all the other PDBs out there, feel free to contact me > > off list. > > You mention MR: Phaser calculates the anisotropy so you can find the value > > in the first lines of the log. > > > > Staraniso or the UCLA server are good to test if you have anisotropy. > > Staraniso has a newer way of dealing with intensity falloff and accounting > > for it. > > > > All the best > > Vincent > > > > > > > > > > On 13/10/2017 10:58, Paul Miller wrote: > >> I had a similar problem to what you describe. In my case the dataset was > >> severely anisotropic (2.6A, 3.4A, 4.6A in each axis). The R factors were > >> stuck similar to yours but the map looked good. I was told by someone with > >> a much better appreciation of the theory than myself that the anisotropy > >> was causing the problem. > >> > >> It would be interesting to know from an expert in anisotropy e.g. the > >> creators of UCLA anisotropy server or Startaniso whether anisotropy can > >> cause this problem and whether there is any way around it. > >> > >> Cheers, Paul > >> > >> Paul Steven Miller (PhD) > >> Postdoctoral Researcher > >> University of Oxford > >> Wellcome Trust Centre for Human Genetics > >> Division of Structural Biology > >> Roosevelt Drive > >> Oxford > >> OX3 7BN > >> > >> > >> Original message > >> > >>> Date: Fri, 13 Oct 2017 10:30:22 +0200 > >>> From: CCP4 bulletin board > >>> (on behalf of Gianluca Cioci > >>> > >>> ) > >>> Subject: [ccp4bb] High R/Rfree after MR > >>> To: > >>> CCP4BB@JISCMAIL.AC.UK > >>> > >>> > >>> Dear All, > >>> I am trying to refine a structure at 3.3A. Model has > >>> 60% identity to the target. Maps look OK (for 3.3A) > >>> and rebuilding in Coot is relatively > >>> straightforward. However, after some rebuilding > >>> cycles the R factors are stuck at 0.37/0.39 > >>> (REFMAC). > >>> XTRIAGE tells me that everything is normal (no twin, > >>> 98% completeness, R=3.5% in the low resolution bin), > >>> perhaps some anisotropy is present. > >>> I have already refined 2 homologous structures at > >>> resolutions going from 3.2 to 3.8 and there were no > >>> problems (final R ~ 0.21/0.24). > >>> An
Re: [ccp4bb] High R/Rfree after MR
Thank to all for the replies ! I have tried the UCLA server using the unmerged intensities from XDS and the verdict is strong anisotropy The Rfactors (after Phaser+Refmac) are only slightly lower for the Corrected. Non Corrected:Corrected R factor 0.3778 R factor 0.3732 R free 0.3871 R free 0.3816 I will make a few cycles of rebuilding and see if it makes any difference... Best regards, GIA Best regards, GIA On Fri, Oct 13, 2017 at 5:11 PM, Randy Read wrote: > Just to add to this point. The MR algorithms in Phaser are now able to > make better use of intensity data, which is particularly important when you > have any very weak data. Having weak data can’t be avoided when you have > serious anisotropy (or tNCS or a combination of the two). Unfortunately, > if you use amplitudes that have been through the French & Wilson (truncate) > algorithm, the real variation in intensity is partially masked because the > posterior amplitude values are computed on the prior assumption that all > the reflections in a resolution shell have the same underlying intensity > distribution. > > The UCLA server actually uses Phaser under the hood — what they add is to > turn the anisotropic B-values into suggested resolution limits in the > different directions. However, I don’t think they allow you yet to submit > intensities, which would be better. > > Best wishes, > > Randy Read > > - > Randy J. Read > Department of Haematology, University of Cambridge > Cambridge Institute for Medical ResearchTel: +44 1223 336500 > Wellcome Trust/MRC Building Fax: +44 1223 336827 > Hills Road > E-mail: rj...@cam.ac.uk > Cambridge CB2 0XY, U.K. > www-structmed.cimr.cam.ac.uk > > > On 13 Oct 2017, at 10:29, vincent Chaptal > wrote: > > > > Dear Gia and Paul, > > > > about anisotropy, one point to keep in mind is that it is not > necessarily linked to the difference in resolution limits. > > In fact I am at the moment working on one of these cases, with extremely > large difference in resolution limits, but relatively low anisotropy. > Anisotropy is more a deviation from "normal" intensity falloff as a > function of resolution. There is a thin difference/relationship between the > two concepts that I think is worth investigating. > > > > we have performed a statistical analysis of this phenomenon and the > paper is in revision at the moment, but if you want to know where your > anisotropy stands in respect to all the other PDBs out there, feel free to > contact me off list. > > You mention MR: Phaser calculates the anisotropy so you can find the > value in the first lines of the log. > > > > Staraniso or the UCLA server are good to test if you have anisotropy. > Staraniso has a newer way of dealing with intensity falloff and accounting > for it. > > > > All the best > > Vincent > > > > > > > > > > On 13/10/2017 10:58, Paul Miller wrote: > >> I had a similar problem to what you describe. In my case the dataset > was severely anisotropic (2.6A, 3.4A, 4.6A in each axis). The R factors > were stuck similar to yours but the map looked good. I was told by someone > with a much better appreciation of the theory than myself that the > anisotropy was causing the problem. > >> > >> It would be interesting to know from an expert in anisotropy e.g. the > creators of UCLA anisotropy server or Startaniso whether anisotropy can > cause this problem and whether there is any way around it. > >> > >> Cheers, Paul > >> > >> Paul Steven Miller (PhD) > >> Postdoctoral Researcher > >> University of Oxford > >> Wellcome Trust Centre for Human Genetics > >> Division of Structural Biology > >> Roosevelt Drive > >> Oxford > >> OX3 7BN > >> > >> > >> Original message > >> > >>> Date: Fri, 13 Oct 2017 10:30:22 +0200 > >>> From: CCP4 bulletin board > >>> (on behalf of Gianluca Cioci < > gianluca.ci...@gmail.com> > >>> ) > >>> Subject: [ccp4bb] High R/Rfree after MR > >>> To: > >>> CCP4BB@JISCMAIL.AC.UK > >>> > >>> > >>> Dear All, > >>> I am trying to refine a structure at 3.3A. Model has > >>> 60% identity to the target. Maps look OK (for 3.3A) > >>> and rebuilding in Coot is relatively > >>> straightforward. However, after some rebuilding > >>> cycles the R factors are stuck at 0.37/0.39 > >>> (REFMAC). > >>> XTRIAGE tells me that everything is normal (no twin, > >>> 98% completeness, R=3.5% in the low resolution bin), > >>> perhaps some anisotropy is present. > >>> I have already refined 2 homologous structures at > >>> resolutions going from 3.2 to 3.8 and there were no > >>> problems (final R ~ 0.21/0.24). > >>> Any advice ? > >>> Thanks, > >>> GIA > >>> > > > > -- > > Vincent Chaptal, PhD > > Institut de Biologie et Chimie des Protéines > > Drug Resistance and Membrane Proteins Laboratory > > 7 passage du Vercors > > 69007 LYON > > FRANCE > > +33 4 37 65 29 01 > > http://www.ibcp.fr > > > > >
Re: [ccp4bb] High R/Rfree after MR
Just to add to this point. The MR algorithms in Phaser are now able to make better use of intensity data, which is particularly important when you have any very weak data. Having weak data can’t be avoided when you have serious anisotropy (or tNCS or a combination of the two). Unfortunately, if you use amplitudes that have been through the French & Wilson (truncate) algorithm, the real variation in intensity is partially masked because the posterior amplitude values are computed on the prior assumption that all the reflections in a resolution shell have the same underlying intensity distribution. The UCLA server actually uses Phaser under the hood — what they add is to turn the anisotropic B-values into suggested resolution limits in the different directions. However, I don’t think they allow you yet to submit intensities, which would be better. Best wishes, Randy Read - Randy J. Read Department of Haematology, University of Cambridge Cambridge Institute for Medical ResearchTel: +44 1223 336500 Wellcome Trust/MRC Building Fax: +44 1223 336827 Hills RoadE-mail: rj...@cam.ac.uk Cambridge CB2 0XY, U.K. www-structmed.cimr.cam.ac.uk > On 13 Oct 2017, at 10:29, vincent Chaptal wrote: > > Dear Gia and Paul, > > about anisotropy, one point to keep in mind is that it is not necessarily > linked to the difference in resolution limits. > In fact I am at the moment working on one of these cases, with extremely > large difference in resolution limits, but relatively low anisotropy. > Anisotropy is more a deviation from "normal" intensity falloff as a function > of resolution. There is a thin difference/relationship between the two > concepts that I think is worth investigating. > > we have performed a statistical analysis of this phenomenon and the paper is > in revision at the moment, but if you want to know where your anisotropy > stands in respect to all the other PDBs out there, feel free to contact me > off list. > You mention MR: Phaser calculates the anisotropy so you can find the value in > the first lines of the log. > > Staraniso or the UCLA server are good to test if you have anisotropy. > Staraniso has a newer way of dealing with intensity falloff and accounting > for it. > > All the best > Vincent > > > > > On 13/10/2017 10:58, Paul Miller wrote: >> I had a similar problem to what you describe. In my case the dataset was >> severely anisotropic (2.6A, 3.4A, 4.6A in each axis). The R factors were >> stuck similar to yours but the map looked good. I was told by someone with a >> much better appreciation of the theory than myself that the anisotropy was >> causing the problem. >> >> It would be interesting to know from an expert in anisotropy e.g. the >> creators of UCLA anisotropy server or Startaniso whether anisotropy can >> cause this problem and whether there is any way around it. >> >> Cheers, Paul >> >> Paul Steven Miller (PhD) >> Postdoctoral Researcher >> University of Oxford >> Wellcome Trust Centre for Human Genetics >> Division of Structural Biology >> Roosevelt Drive >> Oxford >> OX3 7BN >> >> >> Original message >> >>> Date: Fri, 13 Oct 2017 10:30:22 +0200 >>> From: CCP4 bulletin board >>> (on behalf of Gianluca Cioci >>> >>> ) >>> Subject: [ccp4bb] High R/Rfree after MR >>> To: >>> CCP4BB@JISCMAIL.AC.UK >>> >>> >>> Dear All, >>> I am trying to refine a structure at 3.3A. Model has >>> 60% identity to the target. Maps look OK (for 3.3A) >>> and rebuilding in Coot is relatively >>> straightforward. However, after some rebuilding >>> cycles the R factors are stuck at 0.37/0.39 >>> (REFMAC). >>> XTRIAGE tells me that everything is normal (no twin, >>> 98% completeness, R=3.5% in the low resolution bin), >>> perhaps some anisotropy is present. >>> I have already refined 2 homologous structures at >>> resolutions going from 3.2 to 3.8 and there were no >>> problems (final R ~ 0.21/0.24). >>> Any advice ? >>> Thanks, >>> GIA >>> > > -- > Vincent Chaptal, PhD > Institut de Biologie et Chimie des Protéines > Drug Resistance and Membrane Proteins Laboratory > 7 passage du Vercors > 69007 LYON > FRANCE > +33 4 37 65 29 01 > http://www.ibcp.fr > >
[ccp4bb] Problem with ref format (from Rigaku)
Dear all, this is a question about scaling data integrated in CrystalClear (Rigaku data processing gui based on d*trek) in ccp4. Since scaling dtprofit.ref files from different scans is sometimes poor or even failing within CrystalClear (i.e. with dtscaleaverage after merging them), I used to try scaling them with scala. I am facing this problem mainly with high resolution / small molecule data collection, where I need up to 20 scans, each 90-180 degrees, for complete low and high resolution. The procedure, that worked, was using dtrek2scala for each scan (with scan1.ref and output_scan1.head, then a second run of dtrek2scala for scan2.ref and output_scan2.head, etc.) to create scan1.mtz, scan2.mtz, etc. sortmtz with scan1.mtz, scan2.mtz, ... to create a multibatch mtz file scala with the multibatch.mtz file to create the final scaled mtz file Some time ago Rigaku changed the format of the .ref files, so dtrek2scala is not working any more. Is there a possibility to change the new .ref format to the old one? Or can I read the new .ref files in scala or better aimless directly? The alternative to process the images in xds hits a similar problem: The format of the images has also changed and the new .img files (from the Saturn92 detector) are not read anymore (whereas they used to be processable in xds before). Greetings Gottfried Dr. Gottfried Palm Ernst-Moritz-Arndt-Universität Inst. für Biochemie (MNF) Abt. Biochemie I Felix-Hausdorff-Straße 4 17489 Greifswald
[ccp4bb] AW: [ccp4bb] High R/Rfree after MR
Dear GIA, In addition to the anisotropy, I would also check your diffraction images and make sure that there are no (even hardly perceptible) ice rings present. Depending on how the data processing software handled this, they may cause high Rfactors in the range you mentioned. Best, Herman Von: CCP4 bulletin board [mailto:CCP4BB@JISCMAIL.AC.UK] Im Auftrag von Gianluca Cioci Gesendet: Freitag, 13. Oktober 2017 10:30 An: CCP4BB@JISCMAIL.AC.UK Betreff: [EXTERNAL] [ccp4bb] High R/Rfree after MR Dear All, I am trying to refine a structure at 3.3A. Model has 60% identity to the target. Maps look OK (for 3.3A) and rebuilding in Coot is relatively straightforward. However, after some rebuilding cycles the R factors are stuck at 0.37/0.39 (REFMAC). XTRIAGE tells me that everything is normal (no twin, 98% completeness, R=3.5% in the low resolution bin), perhaps some anisotropy is present. I have already refined 2 homologous structures at resolutions going from 3.2 to 3.8 and there were no problems (final R ~ 0.21/0.24). Any advice ? Thanks, GIA
Re: [ccp4bb] High R/Rfree after MR
Dear Gia and Paul, about anisotropy, one point to keep in mind is that it is not necessarily linked to the difference in resolution limits. In fact I am at the moment working on one of these cases, with extremely large difference in resolution limits, but relatively low anisotropy. Anisotropy is more a deviation from "normal" intensity falloff as a function of resolution. There is a thin difference/relationship between the two concepts that I think is worth investigating. we have performed a statistical analysis of this phenomenon and the paper is in revision at the moment, but if you want to know where your anisotropy stands in respect to all the other PDBs out there, feel free to contact me off list. You mention MR: Phaser calculates the anisotropy so you can find the value in the first lines of the log. Staraniso or the UCLA server are good to test if you have anisotropy. Staraniso has a newer way of dealing with intensity falloff and accounting for it. All the best Vincent On 13/10/2017 10:58, Paul Miller wrote: I had a similar problem to what you describe. In my case the dataset was severely anisotropic (2.6A, 3.4A, 4.6A in each axis). The R factors were stuck similar to yours but the map looked good. I was told by someone with a much better appreciation of the theory than myself that the anisotropy was causing the problem. It would be interesting to know from an expert in anisotropy e.g. the creators of UCLA anisotropy server or Startaniso whether anisotropy can cause this problem and whether there is any way around it. Cheers, Paul Paul Steven Miller (PhD) Postdoctoral Researcher University of Oxford Wellcome Trust Centre for Human Genetics Division of Structural Biology Roosevelt Drive Oxford OX3 7BN Original message Date: Fri, 13 Oct 2017 10:30:22 +0200 From: CCP4 bulletin board (on behalf of Gianluca Cioci ) Subject: [ccp4bb] High R/Rfree after MR To: CCP4BB@JISCMAIL.AC.UK Dear All, I am trying to refine a structure at 3.3A. Model has 60% identity to the target. Maps look OK (for 3.3A) and rebuilding in Coot is relatively straightforward. However, after some rebuilding cycles the R factors are stuck at 0.37/0.39 (REFMAC). XTRIAGE tells me that everything is normal (no twin, 98% completeness, R=3.5% in the low resolution bin), perhaps some anisotropy is present. I have already refined 2 homologous structures at resolutions going from 3.2 to 3.8 and there were no problems (final R ~ 0.21/0.24). Any advice ? Thanks, GIA -- Vincent Chaptal, PhD Institut de Biologie et Chimie des Protéines Drug Resistance and Membrane Proteins Laboratory 7 passage du Vercors 69007 LYON FRANCE +33 4 37 65 29 01 http://www.ibcp.fr
Re: [ccp4bb] High R/Rfree after MR
Gianluca, you could become an expert yourself, by trying - the UCLA anisotropy server - STARANISO with these data, and report here what the outcome in each case is - Rwork/Rfree, map appearance, ... My personal experience is that both refmac and phenix.refine deal well with moderate anisotropy (but there is room for improvement, e.g. current phenix.refine does not take the sigmas into account). In cases of severe anisotropy I had good results with STARANISO - better phasing, better maps, better refinement. best, Kay On Fri, 13 Oct 2017 09:58:39 +0100, Paul Miller wrote: >I had a similar problem to what you describe. In my case the dataset was >severely anisotropic (2.6A, 3.4A, 4.6A in each axis). The R factors were stuck >similar to yours but the map looked good. I was told by someone with a much >better appreciation of the theory than myself that the anisotropy was causing >the problem. > >It would be interesting to know from an expert in anisotropy e.g. the creators >of UCLA anisotropy server or Startaniso whether anisotropy can cause this >problem and whether there is any way around it. > >Cheers, Paul > >Paul Steven Miller (PhD) >Postdoctoral Researcher >University of Oxford >Wellcome Trust Centre for Human Genetics >Division of Structural Biology >Roosevelt Drive >Oxford >OX3 7BN > > > Original message >>Date: Fri, 13 Oct 2017 10:30:22 +0200 >>From: CCP4 bulletin board (on behalf of Gianluca >>Cioci ) >>Subject: [ccp4bb] High R/Rfree after MR >>To: CCP4BB@JISCMAIL.AC.UK >> >> Dear All, >> I am trying to refine a structure at 3.3A. Model has >> 60% identity to the target. Maps look OK (for 3.3A) >> and rebuilding in Coot is relatively >> straightforward. However, after some rebuilding >> cycles the R factors are stuck at 0.37/0.39 >> (REFMAC).� >> XTRIAGE tells me that everything is normal (no twin, >> 98% completeness, R=3.5% in the low resolution bin), >> perhaps some anisotropy is present.� >> I have already refined 2 homologous structures at >> resolutions going from 3.2 to 3.8 and there were no >> problems (final R ~ 0.21/0.24).� >> Any advice ? >> Thanks, >> GIA
Re: [ccp4bb] High R/Rfree after MR
I had a similar problem to what you describe. In my case the dataset was severely anisotropic (2.6A, 3.4A, 4.6A in each axis). The R factors were stuck similar to yours but the map looked good. I was told by someone with a much better appreciation of the theory than myself that the anisotropy was causing the problem. It would be interesting to know from an expert in anisotropy e.g. the creators of UCLA anisotropy server or Startaniso whether anisotropy can cause this problem and whether there is any way around it. Cheers, Paul Paul Steven Miller (PhD) Postdoctoral Researcher University of Oxford Wellcome Trust Centre for Human Genetics Division of Structural Biology Roosevelt Drive Oxford OX3 7BN Original message >Date: Fri, 13 Oct 2017 10:30:22 +0200 >From: CCP4 bulletin board (on behalf of Gianluca Cioci >) >Subject: [ccp4bb] High R/Rfree after MR >To: CCP4BB@JISCMAIL.AC.UK > > Dear All, > I am trying to refine a structure at 3.3A. Model has > 60% identity to the target. Maps look OK (for 3.3A) > and rebuilding in Coot is relatively > straightforward. However, after some rebuilding > cycles the R factors are stuck at 0.37/0.39 > (REFMAC). > XTRIAGE tells me that everything is normal (no twin, > 98% completeness, R=3.5% in the low resolution bin), > perhaps some anisotropy is present. > I have already refined 2 homologous structures at > resolutions going from 3.2 to 3.8 and there were no > problems (final R ~ 0.21/0.24). > Any advice ? > Thanks, > GIA
[ccp4bb] High R/Rfree after MR
Dear All, I am trying to refine a structure at 3.3A. Model has 60% identity to the target. Maps look OK (for 3.3A) and rebuilding in Coot is relatively straightforward. However, after some rebuilding cycles the R factors are stuck at 0.37/0.39 (REFMAC). XTRIAGE tells me that everything is normal (no twin, 98% completeness, R=3.5% in the low resolution bin), perhaps some anisotropy is present. I have already refined 2 homologous structures at resolutions going from 3.2 to 3.8 and there were no problems (final R ~ 0.21/0.24). Any advice ? Thanks, GIA
Re: [ccp4bb] AW: [ccp4bb] C-terminal amide
You have labelled the TYC as an L-peptide in the dictionary have you? ( Look at any peptide for the position and format for the label ) I thought REFMAC would then automatically creat a peptide link between residue n and n+1 Eleanor PS and remove the TER record! On 13 October 2017 at 08:08, wrote: > Hi Abhisek (and BB), > > I use the attached cif file. It has an NH2 residue defined as a peptide > and gets automatically linked to the peptide chain in the buster procedure > I use. So if your last residue is Tyr 100, you add NH2 101 as a HETATM in > the peptide chain. > > I have not tested it with Refmac though. > > Best, > Herman > > -Ursprüngliche Nachricht- > Von: CCP4 bulletin board [mailto:CCP4BB@JISCMAIL.AC.UK] Im Auftrag von > Bernhard Lechtenberg > Gesendet: Freitag, 13. Oktober 2017 00:13 > An: CCP4BB@JISCMAIL.AC.UK > Betreff: [EXTERNAL] Re: [ccp4bb] C-terminal amide > > HI Abhisek, > > I had the same problem a few years back. Here is the solution I came up > with thanks to help from the CCP4bb: > > 1) Add pointer atom in Coot NH2 and create link to C-terminal residue > (Coot: Extensions -> Modeling -> Make link) > 2) create cif file with correct link description (see below) > 3) modify LINK record in PDB file to correct residues/ link name (TYR-NH2 > if you use the file below with modifications for your case) > 4) refine in Refmac with cif as LIB in > > Augie Pioszak sent me the cif file copied below. you will have to change > PHE to TYR for your particular case. > > Hope that helps. > > Best, > Bernhard > > --- > > Bernhard, > You need a link record in the pdb file to link the NH2 amino group to your > last peptide residue. When I did this a few years back I had to include a > .cif library file describing the link for use with Refmac. See pdb entry > 3c4m and below for the lib description I used. I still use O, so not sure > how coot handles it, but I would guess it will recognize the NH2 group just > fine. Hope this helps. > Regards, > Augie Pioszak > > # --- LIST OF LINKS --- > # > data_link_list > loop_ > _chem_link.id > _chem_link.comp_id_1 > _chem_link.mod_id_1 > _chem_link.group_comp_1 > _chem_link.comp_id_2 > _chem_link.mod_id_2 > _chem_link.group_comp_2 > _chem_link.name > PHE-NH2 PHE ..NH2 .. > bond_PHE-C_=_NH2-N > # -- > # > # --- DESCRIPTION OF LINKS --- > # > data_link_PHE-NH2 > # > loop_ > _chem_link_bond.link_id > _chem_link_bond.atom_1_comp_id > _chem_link_bond.atom_id_1 > _chem_link_bond.atom_2_comp_id > _chem_link_bond.atom_id_2 > _chem_link_bond.type > _chem_link_bond.value_dist > _chem_link_bond.value_dist_esd > PHE-NH2 1 C 2 N single 1.3290.020 > loop_ > _chem_link_angle.link_id > _chem_link_angle.atom_1_comp_id > _chem_link_angle.atom_id_1 > _chem_link_angle.atom_2_comp_id > _chem_link_angle.atom_id_2 > _chem_link_angle.atom_3_comp_id > _chem_link_angle.atom_id_3 > _chem_link_angle.value_angle > _chem_link_angle.value_angle_esd > PHE-NH2 1 O 1 C 2 N 122.0001.600 > PHE-NH2 1 CA 1 C 2 N 118.2002.000 > loop_ > _chem_link_plane.link_id > _chem_link_plane.plane_id > _chem_link_plane.atom_comp_id > _chem_link_plane.atom_id > _chem_link_plane.dist_esd > PHE-NH2plane11 CA0.020 > PHE-NH2plane11 C 0.020 > PHE-NH2plane11 O 0.020 > PHE-NH2plane12 N 0.020 > # -- > > - > > On Oct 12, 2017, at 2:53 PM, Abhishek Anan > wrote: > > > > Dear all, > > > > I am trying to solve the structure of a peptide with C-terminal amide. > In order to add the amide group to the c-terminal TYR, I substituted TYR > with TYC (tyrosinamide) and created a link between the preceding GLY and > TYC. When refined in refmac, the pdb inserts a TER card between GLY and TYC > and no covalent bond is created between them. How do I fix this? I have > also tried to add NH2 pointer atom to TYR in coot and create a link between > them but in vain. I also imported the NH2.cif file into coot and tried to > make a link but of no use. I also tried to import NH2.cif into Jligand so I > could get a link record but it refuses to open the NH2.cif file. > > > > I would greatly appreciate any help! > > > > Best regards, > > Abhishek >
[ccp4bb] AW: [ccp4bb] C-terminal amide
Hi Abhisek (and BB), I use the attached cif file. It has an NH2 residue defined as a peptide and gets automatically linked to the peptide chain in the buster procedure I use. So if your last residue is Tyr 100, you add NH2 101 as a HETATM in the peptide chain. I have not tested it with Refmac though. Best, Herman -Ursprüngliche Nachricht- Von: CCP4 bulletin board [mailto:CCP4BB@JISCMAIL.AC.UK] Im Auftrag von Bernhard Lechtenberg Gesendet: Freitag, 13. Oktober 2017 00:13 An: CCP4BB@JISCMAIL.AC.UK Betreff: [EXTERNAL] Re: [ccp4bb] C-terminal amide HI Abhisek, I had the same problem a few years back. Here is the solution I came up with thanks to help from the CCP4bb: 1) Add pointer atom in Coot NH2 and create link to C-terminal residue (Coot: Extensions -> Modeling -> Make link) 2) create cif file with correct link description (see below) 3) modify LINK record in PDB file to correct residues/ link name (TYR-NH2 if you use the file below with modifications for your case) 4) refine in Refmac with cif as LIB in Augie Pioszak sent me the cif file copied below. you will have to change PHE to TYR for your particular case. Hope that helps. Best, Bernhard --- Bernhard, You need a link record in the pdb file to link the NH2 amino group to your last peptide residue. When I did this a few years back I had to include a .cif library file describing the link for use with Refmac. See pdb entry 3c4m and below for the lib description I used. I still use O, so not sure how coot handles it, but I would guess it will recognize the NH2 group just fine. Hope this helps. Regards, Augie Pioszak # --- LIST OF LINKS --- # data_link_list loop_ _chem_link.id _chem_link.comp_id_1 _chem_link.mod_id_1 _chem_link.group_comp_1 _chem_link.comp_id_2 _chem_link.mod_id_2 _chem_link.group_comp_2 _chem_link.name PHE-NH2 PHE ..NH2 .. bond_PHE-C_=_NH2-N # -- # # --- DESCRIPTION OF LINKS --- # data_link_PHE-NH2 # loop_ _chem_link_bond.link_id _chem_link_bond.atom_1_comp_id _chem_link_bond.atom_id_1 _chem_link_bond.atom_2_comp_id _chem_link_bond.atom_id_2 _chem_link_bond.type _chem_link_bond.value_dist _chem_link_bond.value_dist_esd PHE-NH2 1 C 2 N single 1.3290.020 loop_ _chem_link_angle.link_id _chem_link_angle.atom_1_comp_id _chem_link_angle.atom_id_1 _chem_link_angle.atom_2_comp_id _chem_link_angle.atom_id_2 _chem_link_angle.atom_3_comp_id _chem_link_angle.atom_id_3 _chem_link_angle.value_angle _chem_link_angle.value_angle_esd PHE-NH2 1 O 1 C 2 N 122.0001.600 PHE-NH2 1 CA 1 C 2 N 118.2002.000 loop_ _chem_link_plane.link_id _chem_link_plane.plane_id _chem_link_plane.atom_comp_id _chem_link_plane.atom_id _chem_link_plane.dist_esd PHE-NH2plane11 CA0.020 PHE-NH2plane11 C 0.020 PHE-NH2plane11 O 0.020 PHE-NH2plane12 N 0.020 # -- - > On Oct 12, 2017, at 2:53 PM, Abhishek Anan wrote: > > Dear all, > > I am trying to solve the structure of a peptide with C-terminal amide. In > order to add the amide group to the c-terminal TYR, I substituted TYR with > TYC (tyrosinamide) and created a link between the preceding GLY and TYC. When > refined in refmac, the pdb inserts a TER card between GLY and TYC and no > covalent bond is created between them. How do I fix this? I have also tried > to add NH2 pointer atom to TYR in coot and create a link between them but in > vain. I also imported the NH2.cif file into coot and tried to make a link but > of no use. I also tried to import NH2.cif into Jligand so I could get a link > record but it refuses to open the NH2.cif file. > > I would greatly appreciate any help! > > Best regards, > Abhishek NH2.cif Description: NH2.cif
