Re: [ccp4bb] How to describe/correct modulated diffraction?
Hi Kevin, A pragmatic way to solve the problem is to index your data in P1 if you have the unmerged data, solve the structure, then revert to the correct space group for refinement. Yes you will have to find 8 copies of your dimer instead of two, but that is not such a big problem, I imagine. You may need to do rigid body refinement too, to be on the safe side, although Phaser does that. I would say your residuals are reasonable for the resolution you quote, but lower numbers would be preferable. The poor density for the second copy would be perhaps due to the approximate location. But if you persist with refinement, you might improve on the situation. Also, the packing pattern might show that the second copy has fewer lattice contacts, causing this 'disorder', something you cannot avoid. The Patterson peak at 0.5, 0, 0.034 means the second copy is shifted by 2.5A from the ideal position along Z (.034 x 147.2= ~5A, which is the separation in Patterson space, but is halved for real space). From the other Patterson peak, the centre of gravity of the first solution would be at 0.25, 0 or 0.5, 0.065 (fractional coordinates), and the second would be at 0.75, 0.5 or 0.0, 0.60. You might see weak fringes in the diffraction pattern if you view it in VIEWHKL, with the l axis in the display plane. The 5A modulation means the weak fringes in the even layers appear around l=30 or 60 (you can't see the 90th order as it is beyond the resolution limit), and in the odd layers they will be at ~15 and ~45 (order 75 might be on the edge of the pattern). Also, the N(z) plot in TRUNCATE should show a distinct shift of the observed curves to the left of the theoretical curves. I had a similar situation way back when, 3DZW, which I never got to publication despite the coordinates release, but the diagnosis followed the above. Good luck. Pierre *** Dr Pierre Rizkallah, Senior Lecturer Structural Biology Institute of Infection & Immunology, Wales Heart Research Institute, School of Medicine, Heath Campus, Cardiff, CF14 4XN email: rizkall...@cardiff.ac.uk http://medicine.cardiff.ac.uk/person/pierre-rizkallah From: CCP4 bulletin board On Behalf Of Dr. Kevin M Jude Sent: Monday, November 13, 2023 10:23 PM To: CCP4BB@JISCMAIL.AC.UK Subject: [ccp4bb] How to describe/correct modulated diffraction? External email to Cardiff University - Take care when replying/opening attachments or links. Nid ebost mewnol o Brifysgol Caerdydd yw hwn - Cymerwch ofal wrth ateb/agor atodiadau neu ddolenni. tl;dr: My data has native Patterson peaks but no tNCS; I can solve the structure and refine to reasonable R/Rfree, but much of the map is poor quality. More details: I have crystallized a heterodimer of Z domains (small three-helix bundles, like pdb id 8DA3) in a few different crystal forms that all diffracted in the 2-2.5 Å resolution range, though in different space groups and different asymmetric unit volumes. I'll focus on the most successful crystal, space group C2221 a = 39.9 b = 74.8 c = 147.2 Å. All of these data sets have native Patterson peaks (for the crystal in question, there are peaks at [0.5, 0, 0.034] and [0.127, 0, 0.131] that are both 23% of the origin height) but tNCS search mode in phaser failed. Turning off tNCS and searching separately for chain A and chain B of the heterodimer, I am able to find two heterodimers - that are not related by simple translation. TFZ for the first heterodimer is 14.6, which decreases to 13.1 after finding the second heterodimer, though LLG increases. Electron density for the first heterodimer (AB) is very good, with easily identifiable side chains. Density for the second heterodimer (CD) is pretty dodgy; though some side chains are identifiable, chain D has tube-like helices. Using NCS restraints, I'm able to refine to Rfree/Rwork 0.27/0.24, but can only place 3 water molecules (with 240 protein residues in the ASU). All of this suggests to me that there must be some kind of modulation of my reflection intensities, though nothing is apparent to me in studying raw or pseudoprecession images. If possible I'd like to figure out how to correct it, but barring that I would be happy to describe it and possibly relate it to my structure. I'm at a loss for the next step, though, and would appreciate any advice from the community on how to approach this. Best wishes to all, Kevin -- Kevin M. Jude, PhD Structural Biology Research Specialist, Garcia Lab Howard Hughes Medical Institute Stanford University School of Medicine Beckman B177, 279 Campus Drive, Stanford CA 94305 To unsubscribe from the CCP4BB list, click the following link: https://www.jiscmail.ac.uk/cgi-bin/WA-JISC.exe?SUBED1=CCP4BB=1 To unsubscribe from the CCP4BB list, click the following link: https://www.jiscmail.ac.uk/cgi-bi
[ccp4bb] PhD opportunity in Virotherapy
To All Aspiring PhD Candidates, Please consult the link https://www.findaphd.com/phds/project/phd-in-cancer-virotherapies-novel-strategies-for-developing-tumour-selective-virotherapies/?p118454 for an opportunity to apply for a PhD training course at Cardiff University School of Medicine. It is a multidisciplinary project, as described on the web page, including Structural Biology. Interested persons should complete and submit the online form after reading all the specifics. Please note that the start date for the post is 1 April 2021, in 7 weeks! Pierre Rizkallah *** Dr Pierre Rizkallah, Senior Lecturer Structural Biology Institute of Infection & Immunology, Sir Geraint Evans Building, School of Medicine, Heath Campus, Cardiff, CF14 4XN email: rizkall...@cardiff.ac.uk<mailto:rizkall...@cardiff.ac.uk>phone: +44 29 2074 2248 http://www.cardiff.ac.uk/people/view/126690-rizkallah-pierre To unsubscribe from the CCP4BB list, click the following link: https://www.jiscmail.ac.uk/cgi-bin/WA-JISC.exe?SUBED1=CCP4BB=1 This message was issued to members of www.jiscmail.ac.uk/CCP4BB, a mailing list hosted by www.jiscmail.ac.uk, terms & conditions are available at https://www.jiscmail.ac.uk/policyandsecurity/
Re: [ccp4bb] number of frames to get a full dataset?
You’re missing out on a grand opportunity for iconoclasm here. Try out ‘Degree of Replication’ or ‘Average Replication’ or ‘Replicate Frequency’. Any other offerings! P. *** Dr Pierre Rizkallah, Senior Lecturer Structural Biology Institute of Infection & Immunology, Sir Geraint Evans Building, School of Medicine, Heath Campus, Cardiff, CF14 4XN email: rizkall...@cardiff.ac.uk<mailto:rizkall...@cardiff.ac.uk>phone: +44 29 2074 2248 http://www.cardiff.ac.uk/people/view/126690-rizkallah-pierre From: CCP4 bulletin board On Behalf Of Bernhard Rupp Sent: 29 June 2020 23:36 To: CCP4BB@JISCMAIL.AC.UK Subject: Re: [ccp4bb] number of frames to get a full dataset? I think it is time to escalate that discussion to crystallographic definition purists like Massimo or to a logical consistency proponent like Ian who abhors definitional vacuum Cheers, BR From: CCP4 bulletin board mailto:CCP4BB@JISCMAIL.AC.UK>> On Behalf Of Andreas Förster Sent: Monday, June 29, 2020 15:24 To: CCP4BB@JISCMAIL.AC.UK<mailto:CCP4BB@JISCMAIL.AC.UK> Subject: Re: [ccp4bb] number of frames to get a full dataset? I like to think that the reflections I carefully measured at high multiplicity are not redundant, which the dictionary on my computer defines as "not or no longer needed or useful; superfluous" and the American Heritage Dictionary as "exceeding what is necessary or natural; superfluous" and "needlessly repetitive; verbose". Please don't use the term Needless repetitivity in your Table 1. It sends the wrong message. Multiplicity is good. All best. Andreas On Tue, Jun 30, 2020 at 12:03 AM James Holton mailto:jmhol...@lbl.gov>> wrote: I have found that the use of "redundancy" vs "multiplicity" correlates very well with the speaker's favorite processing software. The Denzo/HKL program scalepack outputs "redundancy", whereas scala/aimless and other more Europe-centric programs output "multiplicity". At least it is not as bad as "intensity", which is so ambiguous as to be almost useless as a word on its own. -James Holton MAD Scientist On 6/24/2020 10:27 AM, Bernhard Rupp wrote: > Oh, and some of us prefer the word 'multiplicity' ;-0 Hmmm…maybe not. ‘Multiplicity’ in crystallography is context sensitive, and not uniquely defined. It can refer to 1. the position multiplicity (number of equivalent sites per unit cell, aka Wyckoff-Multiplicity), the only (!) cif use of multiplicity 2. the multiplicity of the reflection, which means the superposition of reflections with the same d (mostly powder diffraction) 3. the multiplicity of observations, aka redundancy. While (a) and (b) are clearly defined, (c) is an arbitrary experimental number. How from (a) real space symmetry follows (b) in reciprocal space (including the epsilon zones, another ‘multiplicity’) is explained here https://scripts.iucr.org/cgi-bin/paper?a14080<https://eur03.safelinks.protection.outlook.com/?url=https%3A%2F%2Fscripts.iucr.org%2Fcgi-bin%2Fpaper%3Fa14080=02%7C01%7Crizkallahp%40CARDIFF.AC.UK%7C9d4374a0f1514697edbf08d81c7cdfe1%7Cbdb74b3095684856bdbf06759778fcbc%7C1%7C0%7C637290669953538712=nROKLFzfjRttVaIi8qOBIbY5bB3IGusRudGkb6MX9t0%3D=0> and also on page 306 in BMC. Too much multiplicity might create duplicity… Cheers, BR Jon Cooper On 23 Jun 2020 22:04, "Peat, Tom (Manufacturing, Parkville)" mailto:tom.p...@csiro.au>> wrote: I would just like to point out that for those of us who have worked too many times with P1 or P21 that even 360 degrees will not give you 'super' anomalous differences. I'm not a minimalist when it comes to data- redundancy is a good thing to have. cheers, tom Tom Peat Proteins Group Biomedical Program, CSIRO 343 Royal Parade Parkville, VIC, 3052 +613 9662 7304 +614 57 539 419 tom.p...@csiro.au<mailto:tom.p...@csiro.au> From: CCP4 bulletin board mailto:CCP4BB@JISCMAIL.AC.UK>> on behalf of 0c2488af9525-dmarc-requ...@jiscmail.ac.uk<mailto:0c2488af9525-dmarc-requ...@jiscmail.ac.uk> <0c2488af9525-dmarc-requ...@jiscmail.ac.uk<mailto:0c2488af9525-dmarc-requ...@jiscmail.ac.uk>> Sent: Wednesday, June 24, 2020 1:10 AM To: CCP4BB@JISCMAIL.AC.UK<mailto:CCP4BB@JISCMAIL.AC.UK> mailto:CCP4BB@JISCMAIL.AC.UK>> Subject: Re: [ccp4bb] number of frames to get a full dataset? Someone told me there is a cubic space group where you can get away with something like 11 degrees of data. It would be interesting if that's correct. These minimum ranges for data collection rely on the crystal being pre-oriented, which is unheard-of these days, although they can help if someone is nagging you to get off the beam line or if your diffraction fades quickly. Going for 180 degrees always makes sense for a well-behaved crystal, or 360 degree
Re: [ccp4bb] Powder diffraction database
The database you want is at the International centre for diffraction data, icdd.com , and look in the Powder Diffraction File, PDF . I have never used it myself, and I imagine you need to query it with a diffraction pattern, however it can be parsed. Taking a simile of a pattern extracted from diffraction images, intended for protein crystal diffraction, may not be simple, as there often is preferred orientation, which gives lumps of high intensity around the ring. But it might be sufficient for 'phase recognition'. CDS is intended for curated, refined, single crystal structures, an extremely useful resource, but not for what you have in mind. Good Luck. Pierre Rizkallah *** Dr Pierre Rizkallah, Senior Lecturer Structural Biology Institute of Infection & Immunology, Sir Geraint Evans Building, School of Medicine, Heath Campus, Cardiff, CF14 4XN email: rizkall...@cardiff.ac.uk phone: +44 29 2074 2248 http://www.cardiff.ac.uk/people/view/126690-rizkallah-pierre -Original Message- From: CCP4 bulletin board On Behalf Of Peer Mittl Sent: 02 December 2019 09:51 To: CCP4BB@JISCMAIL.AC.UK Subject: [ccp4bb] Powder diffraction database Could someone please give me some advice on how to query a publicly available powder diffraction database? Upon (protein) crystallization we always get large spherulites of an inorganic compound and I would like to know what it is. It should be possible to use the scattering angles of these spherulites (its definitely not ice) to query a powder diffraction database. But which database (e.g. CSD) and how? All the best, Peer To unsubscribe from the CCP4BB list, click the following link: https://eur03.safelinks.protection.outlook.com/?url=https%3A%2F%2Fwww.jiscmail.ac.uk%2Fcgi-bin%2Fwebadmin%3FSUBED1%3DCCP4BB%26A%3D1data=01%7C01%7Crizkallahp%40CARDIFF.AC.UK%7Cd8f14c3c7d4a4f4eebbb08d7770d325f%7Cbdb74b3095684856bdbf06759778fcbc%7C1sdata=vjw%2BuUs5CCIHiw%2Fkk4lcxvLXsC3GCIpZG8n1P6KuLSU%3Dreserved=0 To unsubscribe from the CCP4BB list, click the following link: https://www.jiscmail.ac.uk/cgi-bin/webadmin?SUBED1=CCP4BB=1
Re: [ccp4bb] TLS parameters
I can vouch for TLS blocks being used by REFMAC and the PDB validation servers, after some frustration I had with a deposition recently: Towards the end of a refinement, I renamed some chains, to make oligomers in the a.u. contiguous in real space. Validation told me the centre of gravity as declared in the header is not the same as that produced from the coordinates. I edited the input TLS file, but it still produced the same outcome. I eventually realised that REFMAC reads the TLS blocks from the input PDB after reading all the other input instructions for the refinement run. This happening last, it overrides the declarations in the input TLS definitions file. In order to get the coordinates and the definitions to match, I had to remove the TLS blocks, produced by an earlier run of REFMAC, from the pdb input file, so that the new definitions can be followed, and appropriate TLS blocks produced. REFMAC would use pre-existing TLS blocks if they are in the PDB file. The mismatch notwithstanding, REFMAC still worked correctly, although the shifts from the group origins would have looked strange if one tries to analyse the TLS motions with TLSANL. I admit, I don't view them. Moral of the story is, be careful when you rename chains at the end of the refinement! Pierre Rizkallah *** Dr Pierre Rizkallah, Senior Lecturer Structural Biology Institute of Infection & Immunology, Sir Geraint Evans Building, School of Medicine, Heath Campus, Cardiff, CF14 4XN email: rizkall...@cardiff.ac.uk phone: +44 29 2074 2248 http://www.cardiff.ac.uk/people/view/126690-rizkallah-pierre -Original Message- From: CCP4 bulletin board On Behalf Of Robbie Joosten Sent: 19 November 2019 15:14 To: CCP4BB@JISCMAIL.AC.UK Subject: Re: [ccp4bb] TLS parameters Hi Eleanor, The blocks are reliably recorded in PDB entries but in some cases the renumbering of residues was not pushed through to TLS groups. Certain selections cannot be captured in the PDB format, for instance the split in main chain and side chain that Refmac allows. Fortunately that feature is hardly used. Parsing TLS records is not straightforward, particularly the sets from Buster suffered a lot from inconsistent manual editing in the early days of TLS refinement. PDB-REDO's extractor does a decent job in getting selections and changing those into Refmac format, but there are definitely cases that it cannot do. We also have a tool that does this for mmCIF files which is not written by me and (therefore) much more sophisticated in handling more complicated cases. Cheers, Robbie > -Original Message- > From: CCP4 bulletin board On Behalf Of Eleanor > Dodson > Sent: Tuesday, November 19, 2019 3:59 PM > To: CCP4BB@JISCMAIL.AC.UK > Subject: [ccp4bb] TLS parameters > > Does anyone know how reliably the different programs record and use > these blocks from the PDB file? > > Eleanor > > > > > To unsubscribe from the CCP4BB list, click the following link: > https://eur03.safelinks.protection.outlook.com/?url=https%3A%2F%2Fwww. > jiscmail.ac.uk%2Fcgi-bin%2Fwebadmin%3FSUBED1%3DCCP4BB%26A%3D1data > =01%7C01%7Crizkallahp%40CARDIFF.AC.UK%7Cf2a9834dd919481f28d508d76d030e > 63%7Cbdb74b3095684856bdbf06759778fcbc%7C1sdata=BMwtQsLcQzHTeZcKqC > Yg5W7MLDM7Phk0MUx8ohylApI%3Dreserved=0 To unsubscribe from the CCP4BB list, click the following link: https://eur03.safelinks.protection.outlook.com/?url=https%3A%2F%2Fwww.jiscmail.ac.uk%2Fcgi-bin%2Fwebadmin%3FSUBED1%3DCCP4BB%26A%3D1data=01%7C01%7Crizkallahp%40CARDIFF.AC.UK%7Cf2a9834dd919481f28d508d76d030e63%7Cbdb74b3095684856bdbf06759778fcbc%7C1sdata=BMwtQsLcQzHTeZcKqCYg5W7MLDM7Phk0MUx8ohylApI%3Dreserved=0 To unsubscribe from the CCP4BB list, click the following link: https://www.jiscmail.ac.uk/cgi-bin/webadmin?SUBED1=CCP4BB=1
Re: [ccp4bb] old data - headers
Hi Dean, Harry, Station 14.1 was also run at 1.488A, partly to maintain the lower energy capacity that disappeared with the closure of 7.2. We are talking here about the last period of SRS Daresbury, in 2005 when staffing and equipment were in 'high flux'. Marccd never went on 14.1, it was ADSC Q4 detector, maybe from the second generation of that particular brand. Note that 2304 pixels times 0.0816 mm per pixel would come to about 188 mm square face of the detector, hence the beam centre, read from the header, being slightly away from 94 x 94 mm. This is certainly fin de siècle feeling! A far cry from where we are now. Please do mention the venerable 14.1 beamline when it comes to depositing your data. Best wishes. Pierre *** Dr Pierre Rizkallah, Senior Lecturer Structural Biology Institute of Infection & Immunology, Sir Geraint Evans Building, School of Medicine, Heath Campus, Cardiff, CF14 4XN email: rizkall...@cardiff.ac.uk phone: +44 29 2074 2248 http://www.cardiff.ac.uk/people/view/126690-rizkallah-pierre -Original Message- From: CCP4 bulletin board On Behalf Of Dean Derbyshire Sent: 31 January 2019 10:21 To: CCP4BB@JISCMAIL.AC.UK Subject: [ccp4bb] FW: [ccp4bb] old data - headers thanks all. I recon I have the ESRF data sorted.. but Daresbury.. am I right in assuming MARCCD or are we going back as far as image plate? here is the image header Harry what do you think HEADER_BYTES= 512; DIM=2; BYTE_ORDER=little_endian; TYPE=unsigned_short; PIXEL_SIZE=0.08160; BIN=none; ADC=slow; DETECTOR_SN=421; DATE=Wed Apr 13 17:59:22 2005; TIME=20.00; DISTANCE=125.000; OSC_RANGE=1.000; PHI=7.000; OSC_START=7.000; AXIS=phi; WAVELENGTH=1.48800; BEAM_CENTER_X=94.700; BEAM_CENTER_Y=96.400; UNIF_PED=1500; SIZE1=2304; SIZE2=2304; CCD_IMAGE_SATURATION=65535; } -Original Message- From: Dean Derbyshire Sent: den 31 januari 2019 10:50 To: 'graeme.win...@diamond.ac.uk' ; 'Luca Jovine' Subject: RE: [ccp4bb] old data ok may have lied.. not just 1 dataset i note. Daresbury 14-1 on 19th April 2005. (I'm assuming MAR image plate but!) ESRF ID23-1 on 4th September 2007. ESRF ID23-1 on 8th November 2007. ESRF ID23-1 on 25th June 2008. ESRF ID23-1 on 11th February2010. I will post the headers in a sec :) -Original Message- From: graeme.win...@diamond.ac.uk [mailto:graeme.win...@diamond.ac.uk] Sent: den 31 januari 2019 10:42 To: Dean Derbyshire Cc: ccp4bb@jiscmail.ac.uk Subject: Re: [ccp4bb] old data Hi Dean Usually there is a serial number buried somewhere in the header - many are text headers though some have TIFF format binary headers. Often a timestamp as well though this is less common From there biosync may help e.g. https://emea01.safelinks.protection.outlook.com/?url=http%3A%2F%2Fbiosync.sbkb.org%2Fbeamlineupdatehistory.jsp%3Fregion%3Deuropean%26synch_id%3Desrf%26bmln_name%3DBM14%26height%3D400%26width%3D600data=01%7C01%7Crizkallahp%40CARDIFF.AC.UK%7Cee66117ff0164e4b1aaf08d68765e7a1%7Cbdb74b3095684856bdbf06759778fcbc%7C1sdata=SErBfjWMz6%2FS%2B7F%2F92N5UPThLB3pjf%2BuaLg5taSabRo%3Dreserved=0 (the list of ESRF MX beamlines is short so should not be too painful) Knowing the format, filename you can probably pin it down or if you share a little more info someone on the BB will know All the best Graeme On 31 Jan 2019, at 09:38, Dean Derbyshire mailto:dean.derbysh...@medivir.com>> wrote: maybe a silly question it there a data base or other way to tell what detector was used to collect historic data. Image header isn’t hugely helpful ESRF is all I know for the source but I’d like to know what the detector was at the time… - I’m talking 2005-2010 Dean Derbyshire Principal Scientist Protein Crystallography [X] Box 1086 SE-141 22 Huddinge SWEDEN Visit: Lunastigen 7 Direct: +46 8 54683219 Mobile: +46731251723 https://emea01.safelinks.protection.outlook.com/?url=www.medivir.comdata=01%7C01%7Crizkallahp%40CARDIFF.AC.UK%7Cee66117ff0164e4b1aaf08d68765e7a1%7Cbdb74b3095684856bdbf06759778fcbc%7C1sdata=ALjE7DR%2FSccWm7lejdFHiMbmCF3fzgLgZHVSUmD6fG0%3Dreserved=0<https://emea01.safelinks.protection.outlook.com/?url=http%3A%2F%2Fwww.medivir.com%2Fdata=01%7C01%7Crizkallahp%40CARDIFF.AC.UK%7Cee66117ff0164e4b1aaf08d68765e7a1%7Cbdb74b3095684856bdbf06759778fcbc%7C1sdata=%2Bic7UXGfsofn01K61C%2BfRRMMEE4ew7QJBVk6uicVGrY%3Dreserved=0> -- This transmission is intended for the person to whom or the entity to which it is addressed and may contain information that is privileged, confidential and exempt from disclosure under applicable law. If you are not the intended recipient, please be notified that any dissemination, distribution or copying is strictly prohibited. If you have received this transmission in error, please notify us immediately. Thank you for your cooperation. To
Re: [ccp4bb] photograph of Enraf-Nonius FAST detector
Hi Harry, Please find attached a picture of the FAST that was installed on its side, at SRS Station 9.6. The polarisation of the synchrotron beam meant we had to move the theta angle in the vertical plane to get to high resolution. Enjoy. Pierre *** Dr Pierre Rizkallah, Senior Lecturer Structural Biology Institute of Infection & Immunology, Sir Geraint Evans Building, School of Medicine, Heath Campus, Cardiff, CF14 4XN email: rizkall...@cardiff.ac.uk<mailto:rizkall...@cardiff.ac.uk>phone: +44 29 2074 2248 http://www.cardiff.ac.uk/people/view/126690-rizkallah-pierre From: CCP4 bulletin board On Behalf Of Harry Powell Sent: 27 November 2018 12:48 To: CCP4BB@JISCMAIL.AC.UK Subject: Re: [ccp4bb] photograph of Enraf-Nonius FAST detector Hi Elspeth That's brilliant - no, it's not too late by any means. Photos of other obsolete detectors are also welcome! On 27 Nov 2018, at 12:40, Elspeth Garman wrote: I have one of the LMB Cambridge one. Can dig it out for you but not till Monday. Is that too late? Elspeth Sent from my iPhone On 27 Nov 2018, at 12:35, Harry Powell <193323b1e616-dmarc-requ...@jiscmail.ac.uk<mailto:193323b1e616-dmarc-requ...@jiscmail.ac.uk>> wrote: Hi I was wondering if anyone out there has a decent-quality photograph of an Enraf-Nonius FAST detector that I could use? I believe that one was installed at SRS Daresbury in 1983... Harry -- Dr Harry Powell To unsubscribe from the CCP4BB list, click the following link: https://www.jiscmail.ac.uk/cgi-bin/webadmin?SUBED1=CCP4BB=1 To unsubscribe from the CCP4BB list, click the following link: https://www.jiscmail.ac.uk/cgi-bin/webadmin?SUBED1=CCP4BB=1
Re: [ccp4bb] ligand binding and crystal form
Hello Veronica, There are 2 aspects to your enquiry: 1- The different number of ligands in your structure could be easily sorted out by a longer soak. You don’t have to stick with 1 minute soak. A day or longer (I’ve heard of a case in which the soak went on for 30 days), would fill up as many sites as can take your ligand. Of course, the soak would have to be neutral to the crystal, i.e. won’t cause any damage. 2- C222 and C2221 are interchangeable if you shift the origin by 0.25in the x direction. You can try reindexing the mtz file to call the space group C222 and solve the structure again, and you will see all structures match up. Or you can simply shift the solution in C2221 by ¼ in X, and you get the same stats, identical. I have done this in the past. But there is one thing you need to do first: You need to permute the C2221 axes so that they are 217.4 354.3 147.0. But this will change the hand, because it isn’t a cyclic permutation. It is like looking down the b-axis with one version or up the b-axis with the other. There is a switch in REINDEX to allow to do that. The b-axis is some 9A longer, but apart from that there will not be much difference. The two cells are the same, effectively. Note that the rogue cell is longer than the other in all 3 dimensions, around 1 or 2 A in x and z, but 9 in y. If you look in the image processing log file, you will see the distance has refined to a slightly different value for this data set, provided you have used the same system for recording the diffraction patterns. The rogue cell could still have a hidden surprise. 3- A proviso for this game of reindexing and origin shift to work, is that the shift should be done in the z-direction, depending on how your molecules pack in the cell. Symmetry and pseudo-symmetry are fascinating games. I hope this gives you a different perspective. Good Luck. Pierre *** Dr Pierre Rizkallah, Senior Lecturer Structural Biology Institute of Infection & Immunology, Sir Geraint Evans Building, School of Medicine, Heath Campus, Cardiff, CF14 4XN email: rizkall...@cardiff.ac.uk<mailto:rizkall...@cardiff.ac.uk>phone: +44 29 2074 2248 http://www.cardiff.ac.uk/people/view/126690-rizkallah-pierre From: CCP4 bulletin board [mailto:CCP4BB@JISCMAIL.AC.UK] On Behalf Of Veronica Fiorentino Sent: 26 October 2016 13:32 To: CCP4BB@JISCMAIL.AC.UK Subject: [ccp4bb] ligand binding and crystal form Hello all, I just solved a NCS-tetrameric (biological assembly is just a dimer) crystal structures with ligand soak (same plate - same conditions). No density for ligand is observed in the first map. In the 2nd, I have 1 ligand bound. In the 3rd, I have 2 ligands bound. Is there any reason for this 'random' behaviour? In addition, I observed just one crystal out of 20 gave a different unit cell. Pointless confirms to me "Best Solution:space group C 2 2 2". REFMAC refinement shows R/Rfree ~ 20/25 % Cell from mtz : 216.5 345.8 145.290.090.090.0 Space group from mtz: number - 21; name - C 2 2 2 All other datasets have: Cell from mtz : 147.0 354.3 217.490.090.090.0 Space group from mtz: number - 20; name - C 2 2 21 I tried re-processing/refining the C2221 dataset in C222 but R/Rfree stays ~45%. Can I also consider the C2221 dataset as a 'different crystal form'? Am I safe? Thank you all, Veronica
Re: [ccp4bb] P3212--1's in Space Group Names?
At the risk of muddying the waters further, the difference between 312 and 321 is that the 2-fold normal to the 3-fold is along the diagonal of the ab face in 312, while it is along the a (or b, by equivalence) in 321. The 1 is definitely no place holder. It has a very significant meaning. The increasing number of apparently redundant numbers in a space group name is not accidental or incremental. It is simply because in e-mail, people could not (in recent history) write the subscripts. So, P3112 used to be written as P 3(1) 1 2, which meant there is a 3sub1 3-fold screw, with a 2-fold normal to it along the diagonal in the ab plane. The round brackets used to save us from confusion, but their use seems to have dropped out from the lexicon. Who'd be a crystallographer today! Pierre Rizkallah -Original Message- From: CCP4 bulletin board [mailto:CCP4BB@JISCMAIL.AC.UK] On Behalf Of Keller, Jacob Sent: 18 February 2015 16:42 To: CCP4BB@JISCMAIL.AC.UK Subject: Re: [ccp4bb] P3212--1's in Space Group Names? Well, I meant no harm to the poor 1 by calling it a placeholder, but that in the case of P3212, the 1 is simply to tell you that there is no rotation about the second axis but is instead about the third. Saying okay, nothing here amounts to being a place-holder to avoid ambiguity in assigning the loci of the rotations. Place-holders are important too, e.g. the 0's in 1000, perhaps. Maybe to be rigorous we should start calling p1 p111? (not really...) JPK -Original Message- From: CCP4 bulletin board [mailto:CCP4BB@JISCMAIL.AC.UK] On Behalf Of Kay Diederichs Sent: Wednesday, February 18, 2015 11:05 AM To: CCP4BB@JISCMAIL.AC.UK Subject: Re: [ccp4bb] P3212--1's in Space Group Names? Hmm, placeholder for me does not seem to emphasize enough the role that this number plays in the space group names. My understanding (but I fail to remember where I read this ...) is that the first number is the order of the rotation (i.e. 6,4,3,2 or 1) of the unique unit cell axis (often the one with the highest symmetry), the second number is the rotation order of a secondary axis, and the third number gives the rotation order of a tertiary axis - which is the third axis in the orthorhombic system, but a diagonal at least in the trigonal and tetragonal (and I think cubic) systems. This makes it clear that each (baseline) letter in the spacegroup name has its specific role, and tells you about the order of the rotation axis. On top of that comes the screw axis information which is much easier to read when using subscripts. But obviously the naming scheme was chosen such that even if screw axes are not indicated with subscripts, the resulting names are unambiguous. best, Kay
Re: [ccp4bb] Cis-peptide bond checking
Hi Tristan, Wouter, Gert and everyone else, I hope that the sudden surge of zeal for eradication of error, re wrongly assigned cis peptides that involve non-proline residues, would not become too great to accept that there are exceptions. I agree, 3% in one entry is probably unjustified, and many reasons could be identified. But there were studies from as long ago as 1991 (Thornton et al), and 1998-1999 (Hilgnefeld et al) and others, where the question of non-proline cis peptides were investigated. A variety of lists were produced, and there are something like 86 (representative) entries with a genuine cis-peptide that is not involving a proline. The CISPEP record in a PDB entry notwithstanding, a map has to justify the assignment. Even when that is true, there ought to be a biological, or at least a biochemical, reason. Indeed there are, in two groups of proteins that I have been involved in, legume lectins and monocot lectins, but I won't go into the justification here. This message is just to emphasise that the crystallographers, the deposition service and the manuscript reviewers ought to be vigilant, to keep our databases clean and fit for purpose. So it is a collective responsibility, and I hope this is borne in mind as the discussion goes on. Thanks for bringing this issue to the fore. Pierre Rizkallah From: CCP4 bulletin board [mailto:CCP4BB@JISCMAIL.AC.UK] On Behalf Of Wouter Touw Sent: 16 February 2015 11:56 To: CCP4BB@JISCMAIL.AC.UK Subject: Re: [ccp4bb] Cis-peptide bond checking Dear Tristan, Thank you for your post of earlier today regarding the problem of cis and trans peptide planes in the PDB. We also realised this problem a while ago and an article describing this problem and a solution is presently under review at Acta Cryst. D. After analysis of the PDB we can state with 95% certainty that ~4600 trans - cis flips in ~2800 entries (and ~70K peptide-plane flips) are needed in the PDB. Around a third of the trans - cis corrections concern non-prolines. We hope to be able to deal with the problem of cis - trans corrections later. In the tradition of our group, the software to detect these flips is already available at swift.cmbi.ru.nl. Hopefully, the referees of our article consider this topic just as important as you and I do :-). Kind regards, Wouter Touw and Gert Vriend On 02/16/2015 10:58 AM, Tristan Croll wrote: Dear all, My apologies for the spam-like nature of my post, but I would like to draw your attention to an important issue (outlined in an upcoming short communication to Acta D, which will appear at doi:10.1107/S1399004715000826 once it's online). At present, neither the structural quality checks in commonly-used crystallography packages nor those run on deposition of a structure to the PDB are flagging the presence of non-proline cis peptide bonds. This has led to the presence of many erroneous cis bonds creeping into the PDB - primarily in low-resolution structures as one would expect, but I have identified clearly erroneous examples in structures with resolutions as high as 1.3 Angstroms. From my analysis, I estimate that a few thousand structures have been affected to some extent, with the worst cases having as high as 3% of their peptide bonds in cis. Particularly if you have published anything 2.5 Angstroms in the past few years, may I gently suggest that you make a quick double-check of your deposited structures? This can be done quickly and simply in Coot (Extensions-Modelling-Residues with Cis peptide bonds). Best regards, Tristan Het Radboudumc staat geregistreerd bij de Kamer van Koophandel in het handelsregister onder nummer 41055629. The Radboud university medical center is listed in the Commercial Register of the Chamber of Commerce under file number 41055629.
Re: [ccp4bb] Sister CCPs
Let me pick up Eleanor#8217;s comment:is there something like a crystallographer today ? Yes, James Holton and countless others. I mean in the true sense ?I think as a #8220;crystallographer#8221; you won#8217;t be able to survive the next decade, you need to diversify your toolset of techniques as pointed out in this articlehttp://www.nature.com/naturejobs/science/articles/10.1038/nj7485-711a Will a spectroscopist survive? Will a biochemist? And I#8217;m not quite sure how software developers see themselves, as I would argue they are typically maybe not doing so much wet lab stuff related to crystallography (I may be wrong here) but rather code these days. What #8220;type#8221; of crystallographer is a software developer ? Perhaps someone like Randy Read and team, or George Sheldrick who started this thread, or Eleanor Dodson whom you are responding to. I think like our beloved crystals #8220;we#8221; come in different flavors. And we need to train the next generation of students with that perspective in mind. Well said. Just my two cents on a snowy day (30cm over night) Jürgen At least you get the snow. We only get the stormy wind and the rain. Pierre Rizkallah (from the windswept Cardiff in Wales) .. Jürgen Bosch Johns Hopkins University Bloomberg School of Public Health Department of Biochemistry Molecular Biology Johns Hopkins Malaria Research Institute 615 North Wolfe Street, W8708 Baltimore, MD 21205 Office: +1-410-614-4742 Lab: +1-410-614-4894 Fax: +1-410-955-2926 http://lupo.jhsph.edu On Feb 13, 2014, at 6:41 AM, Eleanor Dodson eleanor.dod...@york.ac.uk wrote: I agree with Frank - it keeps crystallographers modest to know how challenging wet lab stuff still is.. Eleanor On 12 February 2014 19:23, Robbie Joosten robbie_joos...@hotmail.com wrote: It's not an e-mail bulletin board, but Researchgate seems to be quite popular for wet lab questions. IMO the QA section of the social network is a bit messy. That said, the quality seems to improve gradually. Cheers, Robbie Sent from my Windows Phone Van: Paul Emsley Verzonden: 12-2-2014 19:23 Aan: CCP4BB@JISCMAIL.AC.UK Onderwerp: Re: [ccp4bb] Sister CCPs On 12/02/14 15:59, George Sheldrick wrote: It would be so nice to have a 'sister CCP' for questions aboud wet-lab problems that have nothing to do with CCP4 or crystallographic computing, The is clearly a big need for it, and those of us who try to keep out of wet-labs would not have to wade though it all. FWIW, the remit of CCP4BB, held at jiscmail-central, is describes as: /The CCP4BB mailing list is for discussions on the use of the CCP4 suite, and macromolecular crystallography in general./ Thus wet-lab questions are not off-topic (not that anyone recently described them as such). Having said that, Jiscmail mailing lists are easy to set-up (providing that you can reasonably expect that the mailing list will improve knowledge sharing within the UK centered academic community) and relatively low maintenance. I, for one, would not be entirely unhappy to miss out on questions about lysis. Paul.
Re: [ccp4bb] Superpose, SSM
Dear BB Readers, I have used CCP4 on Windows exclusively for nearly 10 years now. The text editor of choice for PDB files coming out of CCP4 has always been WORDPAD, which comes with Windows, no need to download. NOTEPAD causes me great difficulty with PDB files, probably because of my age, but let's not get hung up on that. I have never been interested enough in chasing up ways of using NOTEPAD properly. In the last couple of months, CCP4 started outputting PDB files that appeared in WORDPAD as one continuous line, i.e. no line feed. This was an INTERMITTENT behaviour. I could not reproduce it to order. These files are readable by NOTEPAD, despite my dislike of this editor. I have not changed anything in my set up or in the CCP4 or Coot installation. So I could blame it on some system updates in WINDOWS that are invisible to me, but then again it could be a fault of something else, perhaps like changing the CRLF combination already mentioned. The files are usually readable in COOT, which reads anything coming out of CCP4, but again output from COOT is occasionally unreadable in CCP4. NOTEPAD readable files are usually OK. Unfortunately, I don't have a recipe for correct action. Having said all that, at the moment output PDB files from CCP4 and COOT appear to be all well behaved in terms of readability, both in the Windows editors and in CCP4. I would like to think this problem has disappeared just as it had appeared suddenly. I shall continue to look out for symptoms of trouble. Pierre *** Dr Pierre Rizkallah, Senior Lecturer in Structural Biology, Wales Heart Research Institute, Heath Campus, Cardiff CF14 4XN email: rizkall...@cardiff.ac.uk phone + 44 29 2074 2248 -CCP4 bulletin board CCP4BB@JISCMAIL.AC.UK wrote: - To: CCP4BB@JISCMAIL.AC.UK From: Robbie Joosten Sent by: CCP4 bulletin board Date: 27-09-2011 05:22 Subject: Re: [ccp4bb] Superpose, SSM One would assume that Windows software would read DOS/Windows type text files... Open the file in Wordpad. Unlike Notepad, it is able to work with Windows and Unix type text files. If you edit something and save the file, it will be in Windows style. If Superpose stops on that, it should really be updated. I'm sure that there are Windows versions op the programs Unix2dos and Dos2unix which were the programs to use to convert one type to the other. You can also use Word to search and replace the linefeeds. Good luck with this very retro problem. Cheers, Robbie Date: Mon, 26 Sep 2011 17:07:50 -0600 From: xtald...@gmail.com Subject: Re: [ccp4bb] Superpose, SSM To: CCP4BB@JISCMAIL.AC.UK I think something in your workflow is inserting dos line feeds (\n\r or \r\n, I can't remember which). If I have guessed correctly, you want to remove those \rs before proceeding (or never let them get in there in the first place). You claim to open it with MS something, which would insert dos line feeds as part of Operation Vendor Lock. Did you happen to save it, perhaps by habit? That would do the trick. It might even do something insidious and insert those linefeeds without your purposefully saving the document. Your best bet to fix the file after corruption is vim (used to be that crystallographers could use real text editors). The command in vim is: :%s/\r//g You might find some third party utility that fixes linefeeds for $30.00 somewhere, if vim is too retro. Otherwise, you may want to start over, skip checking it out in MS something, and go straight to superpose. James On Sep 26, 2011, at 2:51 PM, Matthias Zebisch wrote: Hi again, Thanks for your quick replies but I think I made myself not clear. here is what I'm doing: 1) superpose proteinA.pdb onto proteinB.pdb : works, but gives out proteinA_lsq1.pdb with extra empty lines (not the anisou lines ;o) ) 2) superpose proteinA_lsq1.pdb onto proteinC.pdb : doesnt work because proteinA_lsq1.pdb cannot be read Any ideas? Even if there is some compatibility issue between CCP4 and windows, I guess superpose should be able to read its own files, shouldnt it? Thanks, Matthias On 9/26/2011 9:13 PM, Jacob Keller wrote: I vaguely recall notepad doing something wacky with files in certain cases...why don't you get the excellent text editor NoteTab Light [sic] (I use it all the time--free and works great), then take a look at your files and see whether MS notepad altered the files. JPK On Mon, Sep 26, 2011 at 2:42 PM, Matthias Zebisch matthias.zebi...@bbz.uni-leipzig.de wrote: Dear CCP4 users, I am using the ccp4i version 6.2.0 under windows 7. I've come across a problem with superpose. The outputfile appears to have additional line feeds (see picture) which, however are not seen in the windows notepad. The structure can also be opened in coot and pymol. However, it is not possible to use
Re: [ccp4bb] domain contact surface
You would have to cheat a little bit, and relabel the second domain chain id to something different. Or you can split the pdb file into two. That would still be necessary for using the EBI PISA server at http://www.ebi.ac.uk/msd-srv/prot_int/pistart.html as I have just been doing. In my case I wanted the interface between multi- chain domains, so I had to pretend they were one chain each. It works alright. Pierre Dr. Pierre Rizkallah, Senior Lecturer in Structural Biology, WHRI, School of Medicine, Cardiff University, Heath Park, CF14 4XN email: rizkall...@cf.ac.uk phone + 44 29 2074 2248 Jane Bailey jbailey.x...@googlemail.com 19/02/10 3:44 PM Dear all, I am trying to calculate the domain-domain contact surface from one chain. I see AREAIMOL to only tell you the accessible surface area/residue. Could any other software/webserve could specify residues in the contact surface and calculate the total contact area? Thanks J.
Re: [ccp4bb] domain contact surface
Hello Again, The buried surface area is twice the interface area, because the interface is counted twice when working out the solvent accessible area which disappeared upon interfacing, once on molecule 1 and once on molecule 2. This is a matter of definition. While I prefer to quote the interface area, quite a number of people prefer the buried surface area. Either one should be alright, as long as you say clearly which one you are reporting. Pierre Dr. Pierre Rizkallah, Senior Lecturer in Structural Biology, WHRI, School of Medicine, Cardiff University, Heath Park, CF14 4XN email: rizkall...@cf.ac.uk phone + 44 29 2074 2248 Jane Bailey jbailey.x...@googlemail.com 19/02/10 4:57 PM Ed Pozharski wrote: http://www.ebi.ac.uk/msd-srv/prot_int/pistart.html Try PISA server - it will identify contacts and report total buried surface area per contact. On Fri, 2010-02-19 at 16:42 +0100, Jane Bailey wrote: Dear all, I am trying to calculate the domain-domain contact surface from one chain. I see AREAIMOL to only tell you the accessible surface area/residue. Could any other software/webserve could specify residues in the contact surface and calculate the total contact area? Thanks J. Dear all, Thanks very much for the answers. I managed to get the interface from PISA by assign two chain IDs onto two domains, as suggested by Pierre and Amy. It gave the interface Solvent-accessible area of each domain: 274.2, 312.2, but not the BSA. However I saw it gave the BSA of individual residue from the interface. Do people get the total interface BSA by adding them up? best Jane
Re: [ccp4bb] Refining against images instead of only reflections
Hi Colin, I was slightly disappointed you missed out one thing in your categories: The ice rings. Now if you can model everything, including these, then maybe we can subtract the contribution from these pesky ice rings, and litlerally 'reveal' hidden data. Wouldn't that be nice! As for the hkl multiples, eg 6h 6k 6l, you have to account for extinction, which I believe was coded at some point in the past, but hardly anybody used, as we don't get perfect crystals, or because we simply don't really understand it very well. Just a couple of things to add to your categories. Pierre Dr. Pierre Rizkallah, Senior Lecturer in Structural Biology, WHRI, School of Medicine, Cardiff University, Heath Park, CF14 4XN email: rizkall...@cf.ac.uk phone + 44 29 2074 2248 Colin Nave colin.n...@diamond.ac.uk 23/01/10 11:35 PM Nice overview from Ian - though I think James did make some good points too. I thought it might be helpful to categorise the various contributions to an imperfect diffraction pattern. Categorising things seems to be one of the English (as distinct from Scottish, Irish or Welsh!) diseases. 1. Those that contribute to the structure of a Bragg reflection i) Mosaic structure - limited size and mosaic spread ii) Dislocations, shift and stacking disorders iii) More macroscopic defects giving split spots iv) Unit cell variations (e.g. due to strain on cooling) v) Twinning (?) 2. Diffuse scatter i) Uncorrelated disorder - broad diffuse scatter distributed over image ii) Disorder correlated between cells - sharper diffuse scatter centred on Bragg peaks iii) Related to above inelastic scattering - Brillouin scattering, acoustic scattering, scattering from phonons iv) Compton scattering (essentially elastic but incoherent) v) Fluorescence vi) Disordered material between crystalline blocks but within whole crystal vii) Scatter from mother liquor viii) Scatter from sample mount 3. Instrument effects i) Air scatter ii)Scatter from apertures, poorly mounted beamstop iii) Smearing of spot shapes due to badly matched incident beams, poor detector resolution, too large a rotation range, iv) Detector noise The trouble with categorisation is that one can (Oh no) i) Have multiple categories for the same thing ii) Miss out something important iii) Give impression that categories are distinct when they might merge in to each other. Categorising seagulls (or any species) is an example, perhaps categorising protein folds is too. Not sure about categorising in to English, Scottish etc. All of these flaws will be in the categories above. Despite this, I believe it would help structure determination to have an accurate as possible model of the crystal. This should be coupled with the ability to determine the parameters of the model from the best possible recording instrument. Such a set up would enable better estimates of the intensity of weak Bragg spots in the presence of a high background. There may be an additional gain by exploiting information from the diffuse scatter of the protein. At present, the normal procedure is to treat the background components as the same, have some parameter called mosaicity and use learned profiles derived from nearby stronger spots (ignoring the fact that the intrinsic profiles of a hkl and a 6h 6k 6l reflection will be closely related). The normal procedure is obviously very good but we don't know what we are missing! Any corrections additions to the categories plus other comments welcome Regards Colin -Original Message- From: CCP4 bulletin board [mailto:ccp...@jiscmail.ac.uk] On Behalf Of Ian Tickle Sent: 22 January 2010 10:54 To: CCP4BB@JISCMAIL.AC.UK Subject: Re: [ccp4bb] Refining against images instead of only reflections -Original Message- From: owner-ccp...@jiscmail.ac.uk [mailto:owner-ccp...@jiscmail.ac.uk] On Behalf Of James Holton Sent: 21 January 2010 08:39 To: CCP4BB@jiscmail.ac.uk Subject: Re: [ccp4bb] Refining against images instead of only reflections It is interesting and relevant here I think that if you measure background-subtracted spot intensities you actually are measuring the AVERAGE electron density. Yes, the arithmetic average of all the unit cells in the crystal. It does not matter how any of the vibrations are correlated, it is still just the average (as long as you subtract the background). The diffuse scatter does NOT tell you about the deviations from this average; it tells you how the deviations are correlated from unit cell to unit cell. James, as I've pointed out before this is completely inconsistent with both established DS theory and many experiments performed over the years. If you're simulation is producing this result, then the obvious conclusion is that you're not simulating what you claim to be. I don't know of a single experimental result that supports your claim
Re: [ccp4bb] Coot pudding? (a.k.a N-linked carbohydrate addition)
Hi Damian, I have used both Coot and Refmac for glycan modelling/refinement in the past, last time was 2 or 3 years ago. They both behaved correctly for me. To get the correct behaviour, you must have the correct LINK cards in the header. At the time, the Chemical ID BMA did not exist in the PDB. There was only MAN. Presumably, the PDB had originally decided that the saccahride linkage is a bit like the peptide bond, where it can be one sort or another, so you have to specify it explicitly. But convenience, or avoiding confusion, must have been the reason for the recent creation of BMA. If Coot/Refmac do not have it in their library, well you can add it easily into the correct folder on your disk, by editing the MAN-b-D.cif entry and calling it BMA.cif. Garib told me, when I was grappling with this problem, that Refmac works out from the coordinates which form of the saccahride linkage it is and imposes the correct restraints. Admittedly, there must be room for confusion if modelling is not accurate enough. So BMA is better all round. The branching of the glycan tree does have a good template in PDB entry 1LGB, first deposited in 1994. It has been reviewed to include BMA earlier this year. The branching pattern is correct, so care must be taken to enter the correct numbers in the LINK cards if you are to reproduce a similar branching pattern in your glycan. Incidentally, if you search the PDB for chemical id BMA, there are apparently 36 entries. 1LGB was not among them, and I don't know why. I hope this is useful. Good Luck. Pierre Rizkallah Dr. Pierre Rizkallah, Senior Lecturer in Structural Biology, WHRI, School of Medicine, Cardiff University, Heath Park, CF14 4XN email: rizkall...@cf.ac.uk phone + 44 29 2074 2248 Damian Ekiert dceki...@scripps.edu 22/12/09 5:46 PM For an ideal glycan, you could used a model from a high resolution structure, or something that has been energy minimized, etc. Mostly I find this helps in getting the sugars in about the right place (keeping bond lengths and angles reasonable). I perhaps could stand to fiddle more and maybe I'll look through the updated documentation. Last I tried, the Asn-NAG1 linkage wasn't enforced, with would allow the whole glycan to slide down into the protein, or off into space. I am typically building relatively small glycans (2-5 residues) into ~3A data, so the density itself doesn't keep things in place very well. Regarding BMA vs MAN: When I have tried to used BMA in REFMAC, it doesn't seem to recognize it and requires a library file. But if you use MAN, it adds a MODRES record to the header, enforcing beta-mannose geometry. Not sure if this is just a REFMAC version issue or what. Best, Damian Paul Emsley wrote: If I can chip into this somewhat sacrilegiously-named thread 1) I *would* use real-space refinement :), specifically Sphere Refinement. You can dial down the density weight if needed, of course. 2) the documentation on refining carbohydrates in Coot has recently been updated http://www.biop.ox.ac.uk/coot/doc/coot/Refining-Carbohydrates.html 3) Coot does not (yet) correct chiral centre inversions in glycosidic linkages on refinement 4) or delete the O1s :) Paul. Robbie Joosten wrote: Dear Steve, I would also use Damian's approach, but the sequence of the core should be NAG-NAG-BMA-(MAN)2. This is improtant because the correct stereochemistry restraints for beta-mannose can only be applied when you call the residue BMA. Building carbohydrates also comes with special validation requirements. PDB-care and CARP are both very usefull. Unfortunately, the service is currently down (http://www.dkfz.de/spec/glycosciences.de). Just make sure the links between your carbs are correct and, please, remove the O1 atoms when needed ;) Cheers, Robbie Joosten Date: Mon, 21 Dec 2009 17:48:31 -0800 From: dceki...@scripps.edu Subject: Re: [ccp4bb] Coot pudding? (a.k.a N-linked carbohydrate addition) To: CCP4BB@JISCMAIL.AC.UK Steve, My general strategy is to start with an ideal glycan (an Asn linked to NAG-NAG-(MAN)3 ) and superimpose the Asn on the residue from my protein. Then you can move the whole glycan as a rigid body until the Asn and first NAG are roughly positioned. Then you can tweak any sugars further out on the chain to get them to fit. Unless you have really great density, usually it is best to avoid real pace refine zone. Better to fit the sugars using the manual rigid body fitting tools, do the best you can, then REFMAC usually brings them in OK. I have some models that I could send you if you need them. Best, Damian Ekiert Soisson, Stephen M wrote: Hi everyone- I was searching for some information on what might be the best way to add N-linked sugars in coot, and Google has let me down. Searching
Re: [ccp4bb] Question about merging of data from different crystals
Hi Yuan Cheng, You have received plenty of advice as to how to proceed with your problem. There is still one thing that no one mentioned, and in my opinion, is the simplest way to go about what you want to do. There are plenty of monoclinic cells with a beta that is very close to 90, but when you start with orthorhombic, you don't necessarily know which axis is the unique one. So I would go back to the beginning, and at the autoindexing step, choose the same cell setting for both crystals, whichever space group you choose. You have already identified that a and b had been switched. You don't have to live with that. You can choose a different setting when autoindexing gives you the options. Now, if you are already convinced that it is monoclinic, you still need to know which is the correct b axis, but at least the two data sets should be internally consistent, i.e. R-merge ought to be similar for the overall data set as it is for the individual one, usually an average of the two. Deciding on the correct b axis will have to be done in a different way. Usually MR (I assume you do have a starting model) is very good at selecting the correct setting. You start with your chosen setting, and if that gives you a good solution, then you have got the correct setting very likely. Just to convince yourself, you can try the other setting. It could much better or much worse. It rarely is about the same, because that would require a tetramer in the asymmetric unit, which has a 4-fold local symmetry. I doubt this is what you are dealing with. If I understand the gist of your message you have 2 copies in the a.u. Good Luck. Pierre ** Dr. Pierre Rizkallah, Senior Lecturer in Structural Biology, WHRI, School of Medicine, Academic Avenue, Heath Park, Cardiff CF14 4XN email: rizkall...@cf.ac.uk phone + 44 29 2074 2248 Yuan Cheng ych...@email.unc.edu 22/10/09 9:52 PM Hi everyone, I have some question about the merging of diffraction data from different crystals. I have two (room-temperature) datasets in my hand and each of them can be scaled in P21 space group and the screw axis is along k. However,each dataset only have below 55% completeness. To get better completeness, I am trying to merge this two datasets. The problem is one dataset has cell dimension (80.632,87.085,114.977),(90,90.026,90) and the other one has (85.497,79.857,114.003) (90,90.004,90),. It looks like that the a,b dimensions are switched between these two datasets. Unsurprisingly,the chi^2 is very high when I tried to merge these two datasets in scalepack.I am wondering whether there is any way to reindex these two datasets to make their a/b dimensions match. Any suggestion will be highly appreciated. By the way, as you might notice, the beta angle is pretty close to 90 degree. I scaled the datasets into P212121 or P2221 or P21212 at the beginning,I had no trouble to merge them in these space groups. However,I could not make the Rfree go below 45% during following refinement (2.7 angstrom cutoff). Phenix.xtriage indicated that there might exist twinning but no twin law is given. Then I reindexed the data into P2 and scaled them in P21 SG.Phenix.xtriage indicates there exists a pseudomerohedral twinning operator. When I used the twin law given in phenix.refine,the Rfree could go down to 29%. So I think P21 might be the correct space group. Thanks again for any suggestion. Yuan
Re: [ccp4bb] Summary for Anisotropic Diffraction In Refinement question
Hi Everyone, I echo Andrew's thanks for the summary offered by Justin. I would like to mention another way to trim anisotropic diffraction patterns of the weak patches 'at source', as it were, in MOSFLM, by specifying a sigma cut off applied to each image. from the manual: RESOLUTION [ lowres ] highres [CUTOFF sigcut] The CUTOFF subkeyword allows the resolution limit of reflections written to the MTZ file to be different for each image. The resolution limit is set as that resolution at which I/sigma(I) drops to below sigcut. The I/sigma(I) for fully recorded reflections (if any) is used, otherwise partials. Default sigcut 0.0 I used this option in the past 2 years on one particular data set, which certainly lost all its weak hi-res spots, leaving an ellipsoid of reflections. The merging stats improved significantly, but the completness in the outer shells went down significantly too. I did not compare maps with and without cut-off, in this particular case. If the drop-off is severe, one can imagine the effect on the maps would be streakiness. There is no win-win situation, unfortunately. Some people set the resolution limit to be half-way between that of the best and worst directions, and take everything, weak and strong. It is a sacrifice of some good data for the sake of the overall quality, and produces distortions of its own, maybe without the streaks. Pierre Rizkallah ** Dr. Pierre Rizkallah, Senior Lecturer in Structural Biology, WHRI, School of Medicine, Academic Avenue, Heath Park, Cardiff CF14 4XN email: rizkall...@cf.ac.uk phone + 44 29 2074 2248 A Leslie and...@mrc-lmb.cam.ac.uk 16/09/09 9:24 AM I would like to thank Justin for his summary of this topic, which I'm sure many people found of interest, and is very much in the spirit of the bulletin board. I would just like to correct one factual error, in that it has been possible to specify anisotropic resolution limits to MOSFLM for many years, the appropriate keywords (described in the MOSFLM Help documentation) are: RESOLUTION ANISO 3.5 2.5 2.5 where the three values are the resolution limits along (or close to) a*, b*, c*. Unfortunately this option is not yet available in imosflm. I have not personally used this option and so cannot compare its efficacy relative to integrating isotropically and then applying an anisotropic limit such as Justin describes. Andrew Leslie On 15 Sep 2009, at 21:48, Justin Hall wrote: Dear All; In response to my Anisotropic Diffraction In Refinement, which asked for suggestions for how best to proceed with refinement with an anisotropic data set, I received a large number of responses which overwhelmingly suggested using the UCLA Anisotropy Server (http://www.doe-mbi.ucla.edu/~sawaya/anisoscale/ ). The Anisotripy Server treats scaled/truncated data sets (I used Scala and the old Truncate program). Fo and SigFo are analyzed with respect to resolution in three dimensions and the data treated in three steps: 1) An elliptical resolution boundary is determined and applied. 2) A purely anisotropic B-factor is applied to the Fo and SigFo data to cause the data in all directions to fall off equally. 3) A negative isotropic B-factor is then applied to the structure factors to force the fall-off in the strongest direction to match that of the original data, effectively meaning that the data are not scaled to the mean but the weaker data are scaled up to match the strongest data. Application of a elliptical resolution boundary is justified because the resolution boundary from common integration programs (Denzo and Mosflm for example) is spherical where diffraction for anisotropic data is ellipsoidal. A spherical boundary would result in the inclusion of numerous poorly measured reflections in the higher resolution shells which effectively makes these data more noisy. Imposing an ellipsoidal resolution boundary is equivalent to removing noise from the higher resolution bins and is simply the anisotropic equivalent of the normal resolution limit truncation. However, I was confused by the second and third steps. The second step of application of anisotropic scale factors is appropriate if the refinement program does not include anisotropic scaling in its calculation of Fc, however modern refinement programs do this. Pavel Afonine touched on this in his CCP4BB general posting in response to my original posting where he noted that anisotropic scale factor[s] that [are] part of the total structure factor take care of this (https://www.jiscmail.ac.uk/cgi-bin/webadmin?A2=ind0909L=CCP4BBT=0F=S=P=8362 ). For the third step, applying a negative isotropic B-factor to modify the Fo is equivalent to sharpening the peaks in your maps and this can be useful. However, applying the correction to Fo will also
Re: [ccp4bb] strange pattersons
Hi Francis, The space group will not influence the Patterson, as both C222 and C2221 will become Pmmm in Patterson space. (Aside: You can change the packing pattern from one into the other by shifting the origin by x+0.5, I think, equivalent to adding 0.5 to the x-coordinate. However, you haven't got there yet!). The pattern you show is probably due to the dominance of one or a few reflections that is/are unusually large relative to the others. My guess, from the plot, is H=18, K=20. In a native Patterson, this would be highly unusual, but possible with translational NCS. If this map is some kind of difference Patterson, say anomalous, it is very easy to get such individual reflections that dominate (poor resolution, according to your hint). You have to be careful in selecting the refelctions with these low accuracy measurements. SCALEIT of the data set for analysis would give you a number of measures to select reliable data with: Highest index in each direction, highest and lowest I/sigma (or whatever the coefficient relates to), maximum and minimum numerical values, etc. If all these fail to weed out the erroneous data, you can still exclude specific reflections, provided you know which ones they are. Good Luck. Pierre Rizkallah ** Dr. Pierre Rizkallah, Senior Lecturer in Structural Biology, WHRI, School of Medicine, Academic Avenue, Heath Park, Cardiff CF14 4XN email: rizkall...@cf.ac.uk phone + 44 29 2074 2248 Francis E Reyes francis.re...@colorado.edu 19/08/09 8:40 PM Hi All Im receiving some strange patterns in my pattersons. Space group is C222 with no confidence (due to resolution) of systematic absences to transform to C2221. Thanks! FR
Re: [ccp4bb] shape complementarity calculations
Hi Vaheh, Below is a reply I made to the BB in 2007 after a similar enquiry. I expect the program should still work the same, although I haven't used it in recent releases. You have to run it from line command, as there is no interface for it in CCP4i. If you are running under Windows, as I do, you have to start a DOS prompt window and run it from there. You might even need to specify the full path of the binary executable. You also need to specify the output log file, and you can either type your input directly or put it in a text file. I hope this helps. Pierre p.s. I echo Ed Berry's exhortation: If you have been running MD on your interacting molecules, the SC parameter is probably meaningless. But I am prepared for surprises. * Start of previous Reply *** The SC manual URL http://www.ccp4.ac.uk/dist/html/sc.html does contain all the information you need, but I agree it takes some deciphering. The basic parameters are the selections of two 'molecules' whose interface you want to study. For that purpose, the script or line input is really minimal, such as: molecule 1 zone C 3 C 5 molecule 2 chain D chain E end Here molecule 1 is a number of residues in chain C (zone), and molecule 2 is all of chains D and E. You can include and exclude other atoms in each 'molecule' selection. The instruction 'end' tells the program to start calculating. You can modify the program defaults by adding other instructions for 'dot density', 'probe radius' and others, but I usually run it with all the hard wired defaults. These parameters are largely equivalent to their counterparts in AREAIMOL to determine how accurately you want the interfaces to be calculated. Good Luck. ** End of previous reply *** ** Dr. Pierre Rizkallah, Senior Lecturer in Structural Biology, School of Medicine, Academic Avenue, Heath Park, Cardiff CF14 4XN email: rizkall...@cf.ac.uk phone + 44 29 2074 2248 Oganesyan, Vaheh oganesy...@medimmune.com 03/03/09 10:28 PM Colleagues, Would some one kindly suggest software that calculates shape complementarity of two interacting proteins based on co-crystal structure? I've seen number of reports with sc parameter included but none of those mention how it was done. Among non-runnable programs in CCP4 there is the sc program that indeed does not run. Thanks in advance. ___ Vaheh To the extent this electronic communication or any of its attachments contain information that is not in the public domain, such information is considered by MedImmune to be confidential and proprietary. This communication is expected to be read and/or used only by the individual(s) for whom it is intended. If you have received this electronic communication in error, please reply to the sender advising of the error in transmission and delete the original message and any accompanying documents from your system immediately, without copying, reviewing or otherwise using them for any purpose. Thank you for your cooperation.
[ccp4bb] PhD Studentships at Cardiff Medical School
Dear Bulletin Board Readers, Some PhD studentships have been made available at the Medical Biochemistry and Immunology Dept of the Medical School, Cardiff University, Wales, U.K. There are 2 projects with a structural component, in the T-cell area, detailed at http://www.tcells.org (click on the link near the bottom of the page, where more details are available). The deadline for applications is 13 Mar 2009 (details for the application procedure are also at the above link). Please note that the awards can only cover the costs for U.K. and E.U. nationals only. It would be very much appreciated if this notice is brought to the attention of prospective students. Thank you very much. Pierre Rizkallah ** Dr. Pierre Rizkallah, Senior Lecturer in Structural Biology, School of Medicine, Academic Avenue, Heath Park, Cardiff CF14 4XN email: rizkall...@cf.ac.uk phone + 44 29 2074 2248
