- phenixbb - phenix-online.org

Re: [phenixbb] anomalous difference map calculation
by Alexandra Marques 17 Jun '15

17 Jun '15

Thank you very much for your replies. Here is what I did: 1)I rerun xdsconv again with Friedls law=false in order to get the mtz file with the F(+) and F(-) columns (new.mtz). 2) I used my pdb file from the last refinement together with the new.mtz file as input for “Calculate maps” in Phenix and selected the option for anomalous map. Now I obtained the ANOM PHANOM map. I hope this is correct… In the anomalous map the largest density is in the Mo and Fe atoms. There is no density in the S of SO3 or SO4 groups, so, this will not help me to decide whether there are SO3 groups in the structure (because for the SO4 I have no doubt). However, while searching the mailling list I foud this post http://www.phenix-online.org/pipermail/phenixbb/2009-September/013799.html which worried me. Should I have used from the beginning this “new” mtz file with F+/F- in order to obtain anomalous differences map at each refinement step? Do you think that I should go back, transfer the R-free flags to my new mtz file and repeat all the refinements? Best regards, Alexandra 2015-06-17 16:59 GMT+01:00 Pavel Afonine <pafonine(a)lbl.gov>: > Hi Alexandra, > > to get anomalous difference map you do not need to do anything special: > phenix.refine calculates this map by default as long as your input data are > Fobs(+) and Fobs(-) or Iobs(+) and Iobs(-). > > Refining occupancy of Mo may not be a bad idea given it is likely to be > partially occupied. > > Pavel > > > On 6/17/15 06:04, Alexandra Marques wrote: > > Hi, > > > I am in the last refinement steps of a MR model and I want to calculate > an anomalous difference map essentially to confirm the presence of a > sulfite molecule and to locate vanadium (present in soaking solution). I > read that it is necessary to have a mtz file with anomalous data (i.e. > F+,F- or I+,I-). However, my data was collected at “normal” wavelenght > (0.97) and it was processed with XDS considering Friedls law= true and my > mtz file contain the following columns: H K L FP SIGFP. So, can I still > calculate a anomalous difference map based on my data? > > > Since I also have a Mo atom in the active site can I try to refine its > occupancy by using the option “anomalous groups” in the refinement > strategy? > > > Thank you very much, > Alexandra > > > _______________________________________________ > phenixbb mailing [email protected]://phenix-online.org/mailman/listinfo/phenixbb > Unsubscribe: phenixbb-leave(a)phenix-online.org > > >

3 3

anomalous difference map calculation
by Alexandra Marques 17 Jun '15

17 Jun '15

Hi, I am in the last refinement steps of a MR model and I want to calculate an anomalous difference map essentially to confirm the presence of a sulfite molecule and to locate vanadium (present in soaking solution). I read that it is necessary to have a mtz file with anomalous data (i.e. F+,F- or I+,I-). However, my data was collected at “normal” wavelenght (0.97) and it was processed with XDS considering Friedls law= true and my mtz file contain the following columns: H K L FP SIGFP. So, can I still calculate a anomalous difference map based on my data? Since I also have a Mo atom in the active site can I try to refine its occupancy by using the option “anomalous groups” in the refinement strategy? Thank you very much, Alexandra

4 4

keep torsion restraint on ligand
by CPMAS Chen 17 Jun '15

17 Jun '15

Hi, All Phenix Users, I am fitting 2-bromoethanol to the density map. I want to keep its gauche conformation, so I edited the cif for this ligand to restrain the torsion angle, loop_ _chem_comp_tor.comp_id _chem_comp_tor.id _chem_comp_tor.atom_id_1 _chem_comp_tor.atom_id_2 _chem_comp_tor.atom_id_3 _chem_comp_tor.atom_id_4 _chem_comp_tor.value_angle _chem_comp_tor.value_angle_esd _chem_comp_tor.period BRJ Var_01 BR1 CB CB1 OB1 -60.00 0.0 3 But, after refinement, this angle can be close to 90 or even 120. Even worse is that the fitting at such angles are not better than at -60/60 angle. How could I fix/restrain the angle during refinement? Thanks! Charles *************************************************** Charles Chen Research Associate University of Pittsburgh School of Medicine Department of Anesthesiology ******************************************************

3 3

Re: [phenixbb] anomalous difference map calculation
by Ashok Nayak 17 Jun '15

17 Jun '15

Hello Alexandra, I believe you have to reprocess the data keeping friedel pairs (f+ and f- ) separate to make use of the little most anomalous signal (if any) at your collected wavelength. The best thing would be to collect a high redundant set of peak data (if not all the three data sets including mad remote and inflection data) If you have access to a tunable source and xtals left. Once you process differently, you can do all sorts of substructure search and later refine anomalous scatterers using wavelength, atom type and f ' and f“ values. Sulphur sad is also a good possibility if you have a decent nmbr of sulphurs in your protein. Hope this suffices your query. Regards Ashok Nayak CSIR CDRI, Lucknow, India Alexandra Marques <at.marques(a)fct.unl.pt> wrote: >_______________________________________________ >phenixbb mailing list >phenixbb(a)phenix-online.org >http://phenix-online.org/mailman/listinfo/phenixbb >Unsubscribe: phenixbb-leave(a)phenix-online.org

1 0

phenix refine of my EM data
by Smith Liu 17 Jun '15

17 Jun '15

Dear All, By phenix.real_space_refine, I have fitted the a PDB file to an EM map, with Ramachandran favored around 90% and acceptable level of Ramachandran outlier (1.5%) and rotamer outlier (1.2%). Besides visual checking indicating the fitting of the main chain and sid chains to the dnsity map is acceptable. I hope the refine results can be improved by phenix reciprocal space refine. By phenix.map_to_structure_factors I convert my mrc file to mtz file, with the above mentioned phenix.real_space_refine refined pdb amd mtz files as the input files for the graphical phenix refine, in which I never select real space refine, and I always check xyz coordinates and Group B-factors (sometimes I check xyz coordinates and individual B-factors ). Fot this kind of phenix reciprocal space refine, I find the molprobity anslysis results have improved a little, but I find both the contouring level of 2FOFCWT map and the FOFCWT map is more than thousands of thousands high (and it seems the contouring level of FOFCWT is much higher), and visual checking indicated both the main chain and side chains hardly fit with the density map. But I really find there were publications which did phenix.real_space_refine first, and then process the reciprocal space refine, which get good results as introduced in the publications. Will you please tell me why my phenix reciprocal refine after phenix.real_space_refine did not work? Best regards. Smith

1 0

on phenix.map_to_structure_factors
by Smith Liu 13 Jun '15

13 Jun '15

Dear All For phenix.map_to_structure_factors, it needs to input the resolution d_min. But for an EM map, usually the resolution is the general resolution of the whole map (or to some extent the average resolution of the whole map). Is d_min for phenix.map_to_structure_factors the general EM map resolution, or the best resolution of the whole EM map, which is only for part of the EM map with the best quality? Smith

2 1

on real_space_refine
by Smith Liu 12 Jun '15

12 Jun '15

Dear All, After fitting a PDB into a 3.5-5 A resolution EM map by phenix.real_space_refine, should I still need to do a reciprocal space refine? For crystallography, the reciprocal space is in fact not purely imaginary, it corresponds to the diffaction spot. But for the EM data refinement, what is the reciprocal space refine? And what reciporce spce refine works on for the data refinement? Smith

2 1

refinement of ligands and waters
by Alexandra Marques 10 Jun '15

10 Jun '15

Hi, I still have little experience in the refinement of protein structures. So, I have some (basic) doubts that I would like you to help me please. 1) I have a metal coordinated to a cofactor in the active center of my protein structure. The initial solution provided by phaser/autobuilding already contained the metal+cofactor modelled in the density but with zero occupancy. Then I created all the necessary restraints and cif files with phenix_ready_set and did two cycles of model building/phenix refinement. The refined structures always contained zero occupancy for those atoms. My question is: should I manually change the occupancy for the metal+cofactor to 1? I thought this would be made automatically by phenix because I selected the option “occupancies” and “individual B-factors” in the refinement strategy… 2) When manually placing waters in the density with coot, what is the maximum value of B-factor that is acceptable for a water? I am asking that because I have some water molecules with weak density and consequently B-factors between 45 and 55 but that are involved in interactions with other molecules. Is it better to keep those waters or remove them? Best regards, Alexandra

2 1

does the lastest Phenix only support windows 7
by Xiao Lei 09 Jun '15

09 Jun '15

Hi All, I am a beginner of crystallography and I'd like to install Phenix. I tried to download phenix for my Windows 8.1 system (64bit) but in the download page it says "64-bit (Windows 7 only) ". I do not know if I really have to use Windows 7. Does anyone have experience of installation Phenix on Windows 8.1? Thanks ahead. Xiao

2 2

Berkeley Server Downtime
by Paul Adams 08 Jun '15

08 Jun '15

Dear Colleagues, the CCI group in Berkeley is moving to a new building this week. As a result several structural biology web services will be offline for a short while. These include: - The Phenix web site (http://phenix-online.org/) - The DIALS West web site (http://dials.lbl.gov/) - The CCI web site (http://cci.lbl.gov/) - The Structural Biology Technology Portal (http://technology.sbkb.org/) The servers will be shutdown at 9am Pacific time on Wednesday 10th. Services should resume by the morning of Friday 12th at the latest. During the downtime bug reports and emails requesting help should be queued and received by us once the servers are back online. We greatly appreciate your understanding, and hope that this doesn’t present too much of an inconvenience. Cheers, Paul -- Paul Adams Deputy Division Director, Physical Biosciences Division, Lawrence Berkeley Lab Division Deputy for Biosciences, Advanced Light Source, Lawrence Berkeley Lab Adjunct Professor, Department of Bioengineering, U.C. Berkeley Vice President for Technology, the Joint BioEnergy Institute Laboratory Research Manager, ENIGMA Science Focus Area Building 64, Room 248 Building 80, Room 247 Building 978, Room 4126 Tel: 1-510-486-4225, Fax: 1-510-486-5909 http://cci.lbl.gov/paul Lawrence Berkeley Laboratory 1 Cyclotron Road BLDG 64R0121 Berkeley, CA 94720, USA. Executive Assistant: Louise Benvenue [ LBenvenue(a)lbl.gov ][ 1-510-495-2506 ] --

1 0

no merge original index
by Tarek DawoD 08 Jun '15

08 Jun '15

obtaining unmerged data from HKL2000 when i use "no merge original index" it either does nothing or gives this error ((could not read file /,,,/,,,,//,,///,,,,/.sca header: format is not supported by HKL-2000, File could be corrupted or it may be a result of a "no merge" command)) How could this be fixed?? did somebody else managed to get the unmerged data from HKL2000 or it is problematic? Thank you for your explanation and providing solutionTKDawoD

2 1

problem with B-factor plot within the Structure Comparison tool
by Emilia C. Arturo (Emily) 06 Jun '15

06 Jun '15

Hello All. I am trying to zoom in on a section of the B-factors v. residue plot that is generated using the Structure Comparison tool of the phenix GUI. I zoom in by selecting a narrow residue range. When I do so, the plot does not show all parts of each trace; they are broken up and/or residues are missing that are certainly in the pdb files that I supplied, and that showed up when I displayed a wider residue range. I am using phenix version 1.9-1692 on a Mac (OS X Yosemite). I'd input 21 pdb files. I've done some typical refresher-type things like closed and reopened the project, and restarted phenix (with no rationale, really), but the display problem persists. Does anyone else recognize this problem? Regards, Emily.

2 1

Model bias in MR-SAD
by mohamed noor 05 Jun '15

05 Jun '15

Dear all In the case of MR-SAD, how bad can model bias be? Does it also depend on the data resolution and resolution at which there is still enough anomalous signal? Or can the anomalous signal help in overcoming the bias inherent in the MR model? Thanks.

2 1

R free assignment in thin shells
by mohamed noor 04 Jun '15

04 Jun '15

Dear all I found a not-so-old thread about this topic: http://phenix-online.org/pipermail/phenixbb/2012-January/018259.html. Maybe there has been significant changes in phenix.refine since then. However, I couldn't find a conclusion. In my case, my data extends to 1.4 A with the R/R free being around 23/26 %. There is also pseudo symmetry, so there are 16 NCS copies in a lower symmetry group than what Xtriage suggested. The map looks fine with density for some H atoms even being visible. On what basis should I decide whether to use (or not) thin shell assignment? And what is the cut-off for number of NCS copies? Thanks.

3 2

Problems running phenix.rosetta_refine
by Klose, Thomas 04 Jun '15

04 Jun '15

Dear all, I was trying to set up phenix.rosetta_refine to test it for the refinement of a 4.5Å structure. I installed the latest phenix release and compiled the latest rosetta version (2015.19) using rosetta.build_phenix_interface. However, when I try to run phenix.rosetta_refine I get the following error message: …. Generating 5 models on 4 processors... DEBUG dumping error message ERROR: Rosetta exited with status 1 stderr output: Sorry: Wrong map type requested: %%map_type%% Allowed format is: [p][m]Fo+[q][D]Fc[kick][filled] where [p] and [q] are any numbers (optional), [m] and [D] indicate if the requested map is sigmaa (optional), Fo and Fc are Fobs and Fcalc, [kick] is for Average Kick Map (optional), [filled] is for missing Fobs filled map. Examples: 2mFo-DFc, 3.2Fo-2.3Fc, mFobs-DFcalc_kick, 2mFobs-DFcalc_filled, Fc, 2mFobs-DFcalc_kick_fill, anom, anom_diff, anomalous_difference, Fo ERROR: PhenixInterface::calculateDensityMap() : error writing map ERROR:: Exit from: src/core/scoring/cryst/PhenixInterface.cc line: 1122 This happens with our actual data as well as test data that I downloaded (PDB: 1yip). Any help in solving this problem would be greatly appreciated. Thanks, Thomas

3 4

Help with using FFT map coefficients tool
by Emilia C. Arturo (Emily) 04 Jun '15

04 Jun '15

Hello. I'd like to generate a map appropriate for viewing in PyMOL, using a model and structure factors downloaded from the PDB. But I'm unable to do so, and perhaps you can help me figure out what I might be doing wrong: To generate the map, I did the following using the phenix GUI (version 1.9-1692): 1. I used Reflection tools > Import CIF structure factors to convert the .cif format structure factors for PDB id 1J8U to an mtz file. 2. Then, to configure the job to generate the CCP4 map, I loaded this mtz file, but when I did that I got the following error: "This reflections file does not appear to contain any suitable map coefficients" I went through the same steps again for two other PDB ids, this time generating the mtz file by either letting phenix do it from a PDB id, or by supplying the tool with both a .cif file with structure factors and a pdb model file. I get the same error. Has anyone any advice? Regards, Emily.

3 5

Re: [phenixbb] Changing an amino acid to an unusual amino acid
by Schubert, Carsten [JRDUS] 03 Jun '15

03 Jun '15

Hi Nat, bonds in COOT do not mean much, as they do not translate into bonded interactions in the refinement programs. Phenix for instance tries to figure out the bonding by itself through some distance based heuristics. However in your case it looks as if the CRO residue may not be defined as a polymer, i.e. part of a protein chain in the monomer library you are using. In Windows the monomer library is located in C:\WinCoot\share\coot\lib\data\monomers. Look at the difference in the headers for CYS.cif and CRO.cif or PTR.cif (phosphotyrosine). The aminoacids or modified aminoacids considered part of the peptide chain have all 'peptide' in the header, CRO does not. That is probably the reason why coot does not make any bonds. This would probably cause problems later on with Phenix as well. You could pull out the CRO.cif into your directory, modify it to make it a peptide and read it into coot and phenix during refinement. I would also run it through REEL to make sure the definitions are kosher. HTH Carsten From: Natalia Ketaren [mailto:[email protected]] Sent: Tuesday, June 02, 2015 5:42 PM To: Schubert, Carsten [JRDUS] Subject: Re: Changing an amino acid to an unusual amino acid Hi Carsten, Th residue is a part of the correct chain. It just doesn't make an bonds with the residues on either side of it. Is this common for unusual amino acids when adding to the main chain? Cheers, Nat _____________________________ Natalia E. Ketaren, PhD Postdoctoral Associate Rout Laboratory Box 213 The Rockefeller University 1230 York Avenue New York, NY 10065 Tel: (212) 327-8136 Fax: (212) 327-7193 Natalia.Ketaren(a)rockefeller.edu<mailto:[email protected]> website: http://lab.rockefeller.edu/rout/ NCDIR: http://www.ncdir.org/ On Jun 2, 2015, at 5:26 PM, Schubert, Carsten [JRDUS] <CSCHUBER(a)its.jnj.com<mailto:[email protected]>> wrote: Hi Natalia, if the missing/wrong chain IDs are really the problem you can change them in COOT under Calculate -> Change Chain IDs. The dialog is a bit tricky, since you probably have to use the correct residue range to change an individual residue into an existing chain. In your case I would write out the PDB file and use a text editor to change the chain ID and move the residue into its correct place in the file. HTH Carsten From: phenixbb-bounces(a)phenix-online.org<mailto:[email protected]> [mailto:[email protected]] On Behalf Of Natalia Ketaren Sent: Tuesday, June 02, 2015 5:17 PM To: phenixbb(a)phenix-online.org<mailto:[email protected]> Subject: [phenixbb] Changing an amino acid to an unusual amino acid Hello All, I'm working on a GFP structure and I'm trying to incorporate the CRO (chromophore) residue into the structure using coot. My process is to use Extension->Modelling->Replace residues, where it's suppose to replace a Tyr residue. I type the 3 letter code, CRO, the residue appears, I'm able to real space refine it, but it does not incorporate CRO into the desired chain. Would someone be able to help me figure out what I'm doing wrong? And possibly walk me through a solution? Cheers, Nat _____________________________ Natalia E. Ketaren, PhD Postdoctoral Associate Rout Laboratory Box 213 The Rockefeller University 1230 York Avenue New York, NY 10065 Tel: (212) 327-8136 Fax: (212) 327-7193 Natalia.Ketaren(a)rockefeller.edu<mailto:[email protected]> website: http://lab.rockefeller.edu/rout/ NCDIR: http://www.ncdir.org/

4 3

Negative density
by mohamed noor 03 Jun '15

03 Jun '15

Dear all After refinement, there is a single negative peak in my map where nothing is modeled. What does it mean? Thanks.

3 2

Changing an amino acid to an unusual amino acid
by Natalia Ketaren 02 Jun '15

02 Jun '15

Hello All, I’m working on a GFP structure and I’m trying to incorporate the CRO (chromophore) residue into the structure using coot. My process is to use Extension->Modelling—>Replace residues, where it’s suppose to replace a Tyr residue. I type the 3 letter code, CRO, the residue appears, I’m able to real space refine it, but it does not incorporate CRO into the desired chain. Would someone be able to help me figure out what I’m doing wrong? And possibly walk me through a solution? Cheers, Nat _____________________________ Natalia E. Ketaren, PhD Postdoctoral Associate Rout Laboratory Box 213 The Rockefeller University 1230 York Avenue New York, NY 10065 Tel: (212) 327-8136 Fax: (212) 327-7193 Natalia.Ketaren(a)rockefeller.edu<mailto:[email protected]> website: http://lab.rockefeller.edu/rout/ NCDIR: http://www.ncdir.org/

2 1

Merging reflections
by mohamed noor 02 Jun '15

02 Jun '15

Dear all I have a few native datasets, one anomalous dataset with signal up to 10 A and two other datasets (also collected at the phasing wavelength but no anomalous signal seems to be present) and a sulfur SAD dataset with signal to about 7 A. All the datasets have a resolution of about 2-3 A and processed with xia2/XDS. I couldn't do fluorescence scan as the detector couldn't detect at such a high energy (20 keV). A few questions: 1. Why is there at least some signal from one dataset but not others? The signal should come from a covalently bound ligand, molybdopterin to be precise. 2. Since I(+) should not be equal to I(-) when there is anomalous signal, should I just merge the native dataset together with those collected at phasing wavelength that have no anomalous signal? Or do I merge everything together? 3. Will the anomalous signal to 10 A be useful to do MR-SAD? I could solve the structure with MR but I wanted to be sure my ligand is there. Thanks.

3 4

Allignment of multiple structures
by jens j birktoft 01 Jun '15

01 Jun '15

I apologize if this question this question has been asked before but I still need help finding an answer to the following. I am looking for a program/web-server that will calculate the superposition of multiple structures (non-protein!) Thanks -- +++++++++++++++++++++++++++++++++++++++++++++++++++ Jens J. Birktoft Structural DNA Nanotechnology Department of Chemistry New York University e-mail: jens.knold(a)gmail.com; Phone: 212-749-5057 very slow-mail: 350 Central Park West, Suite 9F, New York, NY 10025 +++++++++++++++++++++++++++++++++++++++++++++++++++

2 1

a question on phenix.real_space_refine
by Smith Liu 01 Jun '15

01 Jun '15

Dear Pavel, By phenix.real_space_refine, I fit a PDB with a 4.0 map. Most of the fitting were nice, but for the Ala residues, it indicates the o of the C=O occupies the density position which should be occupied by the CH3 side chain of the Ala, which also means the CH3 side chain of Ala cannot occupy the density position which clearly should be occupied by the CH3 side chain of Ala. Is any method or restraint to have the CH3 of Ala to ccupy its density position? Smith

1 0

alternate side chains conjugated with water molecules
by Jorge Iulek 31 May '15

31 May '15

Dear all, I have a 1.70 A resolution dataset in which I observe, with certain confidence, that some (few) side chains are alternated with waters. Though managing to tie occupancies of each alternate side chain with a water molecule to give an overall occupancy of 1.00, phenix refine still pushes them (water from side chain) apart as if there were bumping restraints (but their occupancies sum up to no more than 1.00). The closest to this I could find is: http://www.phenix-online.org/pipermail/phenixbb/2009-May/013354.html But I could not find how to turn off specific bumping restraints. I would be thankful if someone could point me to the recipe to do this. Jorge

2 2

Low resolution refinement - rotamers and Ramachandran
by mohamed noor 29 May '15

29 May '15

Dear developers I am refining a low resolution structure (3.8 A). After I fix most of the outliers (red bars) in Coot, phenix.refine is causing my structure to be worse than the start with 13 % Ramachandran outlier and 13 % rotamer outlier. Secondly, when I used phase_and_build to build my model, my R/Rfree is around 30/35 with atoms from some residues missing. When I fill them in with Coot, fix the outliers above and re-refine, R/Rfree jumps to 36/42 and the rmsd angles are too high (2.9). I have already used weight optimization and NCS. Am I doing something wrong? Thanks.

3 5

reference model restraints by segid
by jp d 28 May '15

28 May '15

Hi,I am having trouble getting phenix to used segid's in reference model restraints when my reference and target pdbs both don't use chain ID's.phenix quits with this message: *** Adding Reference Model Restraints *** Sorry: chain ' ' in /home/path/removed/agnref2.pdb contains one or more errant TER cards. Please remove and try again. my definition file looks like this:refinement { main { reference_model_restraints = True } reference_model { file = two.pdb } } refinement.reference_model.reference_group { reference = segid A16S selection = segid A16S } (above about 50 more times) this was on versions 1.9-1692 , dev-2016 and dev-2056 any suggestions ?thanksjpd

4 4