Re: [ccp4bb] SAXS models and data

Hi Gloria, There are repositories of SAXS data. https://www.sasbdb.org/ For example. Hth, Dave Dr David C. Briggs CSci MRSB Principal Laboratory Research Scientist Signalling and Structural Biology Lab The Francis Crick Institute London, UK ==

Re: [ccp4bb] Resolution Discrepancy in Data Set

Hi Liliana, Two things leap out at me when I look at your data summary. (1) Your data probably do not go to 1.77Å. The CC1/2 in your outer shell is below any of the usual thresholds. There are discussions to be had about what the threshold is, but normally CC1/2 values of 0.5 or sometimes 0.3

Re: [ccp4bb] Alternatives to X

9793-7339> | Google Scholar <https://scholar.google.co.uk/citations?user=DRKG5KwJ> From: Robbie Joosten Sent: 06 December 2023 07:56 To: David Briggs ; CCP4BB@JISCMAIL.AC.UK Subject: RE: [ccp4bb] Alternatives to X External Sender: Use cau

Re: [ccp4bb] Alternatives to X

/david_briggs<https://about.me/david_briggs> | OrcID<https://orcid.org/-0002-9793-7339> | Google Scholar <https://scholar.google.co.uk/citations?user=DRKG5KwJ> From: Guillaume Gaullier Sent: 06 December 2023 07:57 To: David

Re: [ccp4bb] Alternatives to X

Hi all, I did a little (and entirely unscientific) test on this with one of our recent papers. Views : Linkedin came out on top. Engagement from other scientists : Mastodon. X didn't really do much, last I checked. There is a structural biology community on Mastodon and there are several

Re: [ccp4bb] [OFF TOPIC] On Ni-NTA resin TEV cleavage

Hi Rafael, For completeness - there are alternatives to imidazole for eluting proteins from Ni-NTA resins: EDTA - (strips Ni2+, and therefore everything bound to the Ni2+) - 50mM should be sufficient, in your favourite buffer/salt system. Obviously not appropriate for metalloproteins. You'll

[ccp4bb] Job Advert: Drug Discovery Project Research Scientist position, Francis Crick Institute, London, UK.

Dear all, We have a position available at the Francis Crick Institute - please share with any soon-to-finish PhD students and postdocs who might be interested: We seek a talented, highly motivated and independent post-doctoral project research scientist to work on a project to design protein

Re: [ccp4bb] AI and cis-peptides

This is very much anecdotal - in my experience, AF2 tends to put the Cys residues in the disulfide bridge adjacent to each other, sometimes with roughly the correct side chain orientation. It doesn't build/refine the S-S bond, becuase I don't think it knows how to. Best, Dave Dr David C.

Re: [ccp4bb] BUSTER on CCP4 Cloud

HI Shawn, the asymmetric unit contents task tells Cloud how many copies of your model exist in the ASU, but not necessarily that they are in the correct position (think, prior to Molecular replacement - no phase information). Depending upon where you are in your workflow, you can run a quick

[ccp4bb] Diamond User Committee nominations

Dear all, (Apologies, this is a message primarily intended for UK-based Diamond Light Source (and potentially eBIC) users, so if that is not you, you can ignore this message) The Diamond User Committee (DUC) is a cross-discipline group of users who provide feedback to the Diamond executive

Re: [ccp4bb] TEV vs HRV3C

Hi Gloria, Both can be made very easily in E.coli. Both are active at 4°C, but especially 3C, I think. I have plasmids for both somewhere in the freezer (you might find someone closer to you who can send HRV3C, but if you cannot, let me know off list). I don't see any particular benefit of one

Re: [ccp4bb] EDR (Sophos) and CCP4 compatibility

Hi Mark, Our IT admin has mandated various AntiVirus software on OSX, including Sophos, and I have never encountered a problem installing and downloading CCP4, Phenix, Coot, Chimera , etc etc HTH, Dave Dr David C. Briggs CSci MRSB Principal Laboratory Research Scientist Signalling and

Re: [ccp4bb] Unidentified electron density

Hi Sayan Looks interesting. My first thought was a metal ion with attendant waters, but it doesn't look quite right for Trigonal co-ordination - maybe trigonal bi-pryamidal with a missing/poorly defined ligand above the plane in image 1? A couple of questions to help refine the search: 1.

Re: [ccp4bb] Regarding R factor value (R Free)

Hi! The R factor describes agreement between the refined model and the processed diffraction data. Without the diffraction data, one cannot calculate an R factor just from atomic positions. To me, it seems like your best bet would be to try and retain the R factor information in the PDB file

Re: [ccp4bb] Has anyone successfully used RoseTTAFold or AF2 to guide crystallization?

Sent: 05 April 2022 14:47 To: CCP4BB@jiscmail.ac.uk Cc: David Briggs Subject: Re: [ccp4bb] Has anyone successfully used RoseTTAFold or AF2 to guide crystallization? External Sender: Use caution. Hi David, do you think that alphafold2 dis/order prediction is better than specific disorder predic

Re: [ccp4bb] Has anyone successfully used RoseTTAFold or AF2 to guide crystallization?

Hi Scott, I've used AF2 order/disorder prediction (based up pLDDT score) to decided upon construct boundaries. We turned a non-expressing construct into a reasonably well expressing construct based on the AF2 prediction. It's part of my construct design process now. HTH, Dave -- Dr David C.

Re: [ccp4bb] mysterious density

MES or HEPES, I reckon. Both of them can lose definition for the electron density in the morpolino/piperazine 6-membered ring due to flexibility between the ring and the sulphonic acid group when - leading to the tadpole-like density you describe. HTH, Dave -- Dr David C. Briggs CSci

Re: [ccp4bb] off-topic: glycans

Hi Sam, GlycoMod from Expasy sounds like it might do what you want to do. https://web.expasy.org/glycomod/ D -- Dr David C. Briggs Senior Laboratory Research Scientist Signalling and Structural Biology Lab The Francis Crick Institute London, UK == Diamond User Committee (MX) CCP4 WG2

Re: [ccp4bb] Eukaryotic protein expression

Hi Dhiraj, For transient expression in HEK293 cells in suspension, I generally use pcDNA3.1 which also uses a CMV promoter. Yields range from nothing to 50mg/l, depending entirely on what it is you're making. I would suggest that maybe the promoter is not at fault, and perhaps the product is.

Re: [ccp4bb] missing covalent bond between asparagine and N-acetylglucosamine using coot carbohydrate module and phenix.refine

Hi Jiang Xu, (Phenixbb added as I suspect this is Phenix not coot that has caused your issue) Did you check the "add hydrogens" button in Phenix.refine? Sometimes this can erroneously add a second H to the ND of glycosylated Asn residues. (I assume this is because bond distances in the input

Re: [ccp4bb] help needed with link description

Hi Gerlind, Rob Nicholls gave a talk on how to deal with covalent linkages using AceDrg at CCP4 study weekend in 2020. https://youtu.be/p4oTJ0bjD3M And Paul Emsley has written up a guide about it... https://pemsley.github.io/coot/blog/2020/06/30/make-a-link.html Hopefully these will be

Re: [ccp4bb] Co crystalization with less soluble ligand.

These are my go-to review articles for ideas for getting a ligand into a crystal. https://scripts.iucr.org/cgi-bin/paper?S0907444906047020 https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5297911/ I have also seen a talk where someone successfully dissolved their water-insoluble ligand in

Re: [ccp4bb] Fluka-branded PEG 3000

Hi Galen, Is it possibly the age of the original material, rather than the batch or brand? PEG does oxidise over time. Best, Dave -- Dr David C. Briggs Senior Laboratory Research Scientist Signalling and Structural Biology Lab The Francis Crick Institute London, UK == about.me/david_briggs

Re: [ccp4bb] Co-crystallization and thermal shift assay

Hi Saif, Whilst in very general terms, ∆Tm does correlate with binding affinity (there will of course always be exceptions to this rule), I don't think there is a cutoff beyond which you know that co-crystallisation is feasible. The degree of stabilisation will depend very much on the system

Re: [ccp4bb] Contagious, Self-Distributing "Vaccines?"

I think it is possible, but the reason it hasn't been done and will probably never be done is that informed consent is an important part of clinical practice. D -- Dr David C. Briggs Senior Laboratory Research Scientist Signalling and Structural Biology Lab The Francis Crick Institute London,

Re: [ccp4bb] AlphaFold: more thinking and less pipetting (?)

A quick note regarding the code that Deepmind released for CASP13 (2018). It bears the rather important caveat that: "This code can't be used to predict structure of an arbitrary protein sequence. It can be used to predict structure only on the CASP13 dataset (links below)." Source:

[ccp4bb] Diamond-II MX beamlines letters of support

Dear all, (Apologies for the slightly off-topic cross-post - hopefully, this will be of interest to all MX practitioners, not just Diamond light source users.) As Diamond User Committee MX representatives, we'd just like to remind you that the deadline for letters of support for Diamond-II

Re: [ccp4bb] File System Problems

Hi Sam, we had a similar issue with CCP4I2 on Linux in the last couple of weeks. Installing the very latest updates made the problem go away for us. I don't know if this Linux issue would also crop up in Windows, but what you describe seems similar - try to open a file selection dialogue and

Re: [ccp4bb] electron density close Histidine side chain

From: samer halabi Sent: 21 July 2020 11:08 To: ccp4bb@jiscmail.ac.uk ; David Briggs Subject: Re: [ccp4bb] electron density close Histidine side chain Hello, Thank you for your kind reply. If the distances are still less than 2.2Å, would coot and Refmac still consider

Re: [ccp4bb] electron density close Histidine side chain

Hi Samer, Did you use a His tag/Ni-NTA during purification? Sometimes Ni2+ ions leach off the Ni-NTA - maybe the two "ears" are accompanying waters? Ni-His co-ordination distance is pretty short (2-2.2Å - table 3 https://www.ncbi.nlm.nih.gov/pmc/articles/PMC2872550/#!po=0.69) and might

Re: [ccp4bb] Commercial source of His-tagged protein

Hi Mark, NEB

Re: [ccp4bb] Molecular replacement problem

Hi Robert, Have you tried lower symmetry spacegroups? Maybe your crystal is 'almost-but-not-quite' orthorhombic and is in fact monoclinic, pretending to be orthorhombic. Zanuda can do this for you. https://www.ccp4.ac.uk/newsletters/newsletter48/articles/Zanuda/zanuda.html Good luck, Dave

Re: [ccp4bb] disinfecting keyboards

Have you considered autoclaving the keyboards in between users? Or maybe autoclave the users? That'll work, right? -- Dr David C. Briggs Senior Laboratory Research Scientist Signalling and Structural Biology Lab The Francis Crick Institute London, UK == about.me/david_briggs

Re: [ccp4bb] CCP4BB vs COVID19

For general interest, The Francis Crick institute in London is offering facilities to public health England, and over 300 Crick scientists have volunteered to help with testing. https://www.crick.ac.uk/news/2020-03-19_francis-crick-institute-offers-assistance-in-emergency-coronavirus-testing

Re: [ccp4bb] Red trapezoids in Coot

Hi Joel, The red trapezoids are indicative of cis-peptide bonds that are exceptionally rare, and usually indicate incorrect tracing of the backbone (unless you have good enough data to unambiguously decide that a cis-peptide bond is correct). I'm afraid I don't know how to get rid of them,

Re: [ccp4bb] protein expression in human cells

7:10:06 AM To: David Briggs ; CCP4BB@JISCMAIL.AC.UK Subject: RE: [ccp4bb] protein expression in human cells Dear colleagues, I wonder if there were a bit less controversial possibility. No matter if that was less efficient. Would there be an option of using human cell lines that do not origin f

Re: [ccp4bb] protein expression in human cells

Hi Gloria, Another vote here for HEK 293 Expi or Freestyle. The off-the-shelf transfection reagents are super expensive, but I make my own (happy to share protocols with anyone interested) and we normally get a few mgs of protein per litre of suspension culture -- certainly enough for

Re: [ccp4bb] Optimization from needle shaped crystals

4. Matrix microseeding. Make a seed stock from these crystals and then re-run your primary screens. https://www.ncbi.nlm.nih.gov/m/pubmed/25195878/ -- Dr David C. Briggs Senior Laboratory Research Scientist Signalling and Structural Biology Lab The Francis Crick Institute London, UK ==

Re: [ccp4bb] Another difficult MR case

Following on from Ivan's suggestion, SIMBAD might he worth a shot. https://journals.iucr.org/d/issues/2018/07/00/rr5159/ The other thing you might try is handing the MR phases to the density modifying and autobuilding program of your choice, increasing the number of cycles by

Re: [ccp4bb] SEC and MALS

Natesh, Did you use an experimentally determined dn/dc value? Is it a glycoprotein, or does it have some other modification? Default dn/dc values are not appropriate for glycoproteins, and will give rise to incorrect Mwt values. Dave -- Dr David C. Briggs Senior Laboratory Research Scientist

Re: [ccp4bb] Problem in real space - please sign & invite other scientists to sign this letter

Dear Daniel, I could not disagree with you more. As scientists it falls to us to support our colleagues (from whatever other discipline) when they provide evidence that has profound ramifications for humanity. Would we hesitate to stand by colleagues who (for example) demonstrate the efficacy

Re: [ccp4bb] Where is M.Harding's website for metal coordination geometry

In addition to MetalPDB there is also Check My Metal. https://csgid.org/metal_sites HTH, Dave From: CCP4 bulletin board on behalf of Eleanor Dodson <176a9d5ebad7-dmarc-requ...@jiscmail.ac.uk> Sent: 07 May 2019

Re: [ccp4bb] analytical gel filtration columns

Hi Mohinder, Have you tried SEC columns like Sephacryl S-400 or CL-4B? They have a higher fractionation range than Superose 6 HTH, Dave From: CCP4 bulletin board on behalf of Mohinder Pal Sent: 17 April 2019 16:10 To: CCP4BB@JISCMAIL.AC.UK Subject: Re:

Re: [ccp4bb] pseudo internal symmetry

Hi, This doesn't answer your question directly, but I'd be tempted to reduce the symmetry and re-refine in a space group lacking that 2-fold axis. I'd hope that the scaling in the lower space group would give you better stats as well, but maybe the differences in your case are too subtle? I

[ccp4bb]

Hi, In Coot 0.8.9.1 Edit -> Preferences -> Others -> Fonts HTH, Dave From: CCP4 bulletin board on behalf of Sanaz Asadollahpour Sent: 12 February 2019 08:22 To: CCP4BB@JISCMAIL.AC.UK Subject: [ccp4bb] Dear All, How can i change the font size of atom labels

Re: [ccp4bb] suggestion of crystallization optimization

Hi Liuqing, 1) When you say 8Å diffraction - did you test the crystals at room temperature? 2) Change the construct. Trim loops and termini, (re)move tags, etc. HTH, Dave From: CCP4 bulletin board on behalf of Liuqing Chen <519198...@163.com> Sent: 04

[ccp4bb] Postdoc Position available - Signalling and Structural Biology Lab, the Francis Crick Institute,

Dear all, A four-year postdoc position is available in the Signalling and Structural Biology Laboratory at the Francis Crick Institute, London, UK, focusing on cryoEM / X-ray crystallography of Rho-dependent protein kinases assemblies. Candidates with experience in biochemistry and/or

Re: [ccp4bb] Protein Ligand interaction using SPR technique

gt; -------- > Praveen > -- [image: --] David Briggs PhD [image: https://]about.me/david_briggs <https://about.me/david_briggs?promo=email_sig_source=email_sig_medium=email_sig_campaign=external_links>

Re: [ccp4bb] protein precipitation reg

eta-mercaptoethanol and 0.5% triton x 100 for lysis, the same buffer with > 20-30mM imidazole for washing and 300mM imidazole for eluting the proteins. > > > > Thank you > > Regards > > Akila > > > > -- > > Akilandeswari G > > > -- [image: --] David Briggs PhD [image: https://]about.me/david_briggs <https://about.me/david_briggs?promo=email_sig_source=email_sig_medium=email_sig_campaign=external_links>

Re: [ccp4bb] Unknown blob extended from catalytic serine residue

with this matter. > > > > Thanks and regards, > > Sharifah Nur Hidayah > Universiti Putra Malaysia, > Malaysia > > > > > > -- > Kevin Jin > > Sharing knowledge each other is always very joyful.. > > Website: http://www.jinkai.org/ > > -- [image: --] David Briggs PhD [image: https://]about.me/david_briggs <https://about.me/david_briggs?promo=email_sig_source=email_sig_medium=email_sig_campaign=external_links>

[ccp4bb] Coot crashes on starting baton mode.

coot-exe: "/Applications/ccp4-7.0/libexec/coot-bin" coot-version: /Applications/ccp4-7.0/libexec/coot-bin platform: /usr/bin/uname core: #f -- [image: --] David Briggs PhD [image: https://]about.me/david_briggs <https://about.me/david_briggs?promo=email_sig_source=email_sig_me

Re: [ccp4bb] Nitrate versus Carbonate

; Chairman of European Crystallographic Association SIG9 (Crystallographic > Computing) > > > > > > > > > > > > > > > -- > Post-Doctoral Research Associate > Texas A University > Dept. of Biochemistry and Biophysics > Interdisciplinary Life Sciences Building > 301 Old Main Drive, Lab 2138 > College Station, Texas 77843 > -- [image: --] David Briggs PhD [image: https://]about.me/david_briggs <https://about.me/david_briggs?promo=email_sig_source=email_sig_medium=email_sig_campaign=external_links>

Re: [ccp4bb] FW: [ccp4bb] Fwd: [ccp4bb] just out of totally idle curiosity ...

Particularly if you are no longer able see that the pursuit of a morally > upright society is actually a good and worthy goal. > > Best, > John > -- [image: --] David Briggs PhD [image: https://]about.me/david_briggs <https://about.me/david_briggs?promo=email_sig_source=email_sig_medium=email_sig_campaign=external_links>

Re: [ccp4bb] just out of totally idle curiosity ...

ochemistry and Molecular Biology > Professor, Department of Chemistry and Biochemistry > and The Center for the Molecular Biology of RNA > University of California at Santa Cruz > Santa Cruz, California 95064 > USA > > http://scottlab.ucsc.edu > > > > > -- > Rica

Re: [ccp4bb] OT: Lunch with the FT: Martin Shkreli

ography, > it’s beautiful,” he says. " > > Google-cache: > > http://webcache.googleusercontent.com/search?q=cache:ee0nCvtuV7kJ:https://www.ft.com/content/693ad306-9b6d-11e6-8f9b-70e3cabccfae > > Reference: https://en.wikipedia.org/wiki/Martin_Shkreli > -- [image: --] David Bri

Re: [ccp4bb] Two SGs in one droplet?

and thanks a lot in advance for your input. > > Regards > > Sam Tang > Biochemistry Programme, School of Life Sciences, CUHK > > -- [image: --] David Briggs PhD [image: https://]about.me/david_briggs <https://about.me/david_briggs?promo=email_sig_source=email_sig_medium=email_sig_campaign=external_links>

Re: [ccp4bb] metal electron density detection

< 0d48627c03ca-dmarc-requ...@jiscmail.ac.uk> wrote: Hi all, Is there any program (like ARP/wARP) available to locate the correct metal atom automatically in the electron density ? best, Ansuman -- [image: --] David Briggs PhD [image: https://]about.me/david_briggs <https:/

Re: [ccp4bb] Ambiguous metal ion at active site.

look at the co-ordination geometry. Chimera has a fantastic tool for doing this (just ctrl-double click on the metal ion and select co-ordination geometry). HTH, Dave *rough, but better than nothing. [image: David Briggs on about.me] David Briggs about.me/david_briggs http://about.me

Re: [ccp4bb] How to fit BioSAXS shape to the Structure

/supcomb.html [image: David Briggs on about.me] David Briggs about.me/david_briggs http://about.me/david_briggs On 26 June 2015 at 12:04, Reza Khayat rkha...@ccny.cuny.edu wrote: Hi, Follow up question on SAXS. Does SAXS have an enantiomer problem like electron microscopy? In other words, does

Re: [ccp4bb] How to fit BioSAXS shape to the Structure

SASTBX has an online tool for achieving this: http://sastbx.als.lbl.gov/cgi-bin/superpose.html [image: David Briggs on about.me] David Briggs about.me/david_briggs http://about.me/david_briggs On 26 June 2015 at 11:39, Ashok Nayak ashokgocrac...@gmail.com wrote: Dear Weifei, It can also

Re: [ccp4bb] cryo condition

If in doubt, try dragging the crystal through a 1:1 mix of Paratone-N and Mineral oil until most or all the mother liquor from surrounding the crystal has been removed. There are more tips here: http://hamptonresearch.com/tip_detail.aspx?id=99 Good luck! D On Mon, 4 May 2015 19:45 Faisal

Re: [ccp4bb] Thin plate crystals

Hi, In my experience, additive screens (e.g Hampton's) can change crystal morphology. You could also re-screen for new conditions either using matrix micro seeding, or change the protein buffer. Perhaps adding a ligand or a component from your current crystallisation conditions to your protein

Re: [ccp4bb] Strange Ancient Diffraction Pattern...

This looks like a tough cookie. IMO you'd be crackers to persist with this crystal form. It's certainly not going to be a piece of cake. Dr David C Briggs PhD http://about.me/david_briggs On 1 Apr 2015 12:07, Keller, Jacob kell...@janelia.hhmi.org wrote: Can anyone index this? It's got mostly

Re: [ccp4bb] how to reduce protein solubility

Hi Francesca, Try some zinc (1mM + ) in your protein buffer? Zinc tends to make a lot of things less soluble in my hands. Dave On Tue, 17 Feb 2015 04:34 Mattiroli,Francesca francesca.mattir...@colostate.edu wrote: Hi all, I am struggling with a protein complex that is too soluble. I have

Re: [ccp4bb] Absence of contact between layers in a crystal

Haven'tthat paper and the associated structure been retracted? http://www.nature.com/news/2009/091222/full/462970a.html There was a huge scandal when it was discovered that Krishna Murthy had falsified data, including the structure you refer to. See

[ccp4bb] X-ray crystallography facility manager position available at Imperial College London

Dear all, A position has come available at Imperial for an X-ray facility manager position. Responsibilities will include maintenance and training for our local X-ray diffraction source and robotic crystallisation facilities, as well as management of BAG time at the Diamond Light Source. Please

Re: [ccp4bb] secondary structure problem...

Mintu, Can you be a little more specific about what your problem is? Are you worried about the rfactors (they don't seem too bad) or the lack of secondary structure? Regarding the secondary structure - some proteins just don't have much! As long as the fit to density and geometry are good, I

[ccp4bb] Protein crystallography facility manager position available at Imperial College London

Dear all, A crystallography facility manager position has become available at Imperial College London. To apply, p lease go to the Imperial jobs website http:// http://www.imperial.ac.uk/job-applicants/www.imperial.ac.uk http://www.imperial.ac.uk/job-applicants//

Re: [ccp4bb] asymmetric homotrimer in the asu

Hi Hay, I think SAXS should be more than capable of discriminating between a 12.5 kDa monomer vs ~37.5 kDa trimer. Lysozyme is a useful standard used in SAXS (as with most structural biology!), and Lysozyme is only slightly larger than your proteins. Cheers, Dave On Fri Dec 12 2014 at 5:13:26

Re: [ccp4bb] protein forming dimers

Ultracentrifugation... SEC with a suitable column for your predicted protein size would probably be a first thing to try. If you have SEC with in-line MALLS, even better. HTH, Dave [image: David Briggs on about.me] David Briggs about.me/david_briggs http://about.me/david_briggs On 20 October 2014 12

Re: [ccp4bb] 3 letter code

When I google 3 letter code for p nitrophenyl phosphate the second site listed is: http://www.ebi.ac.uk/pdbe-srv/pdbechem/chemicalCompound/complete/4NP Which looks like it might contain the answer you seek. HTH, Dave [image: David Briggs on about.me] David Briggs about.me/david_briggs

Re: [ccp4bb] is small dialysis tubes reusable??

. HTH, Dave [image: David Briggs on about.me] David Briggs about.me/david_briggs http://about.me/david_briggs On 26 September 2014 15:51, Manjula Ramu manjula@gmail.com wrote: Hi all, I use Slide-A-Lyzer MINI Dialysis Devices from Thermo for dialysis of small protein volumes

Re: [ccp4bb] off topic

: David Briggs on about.me] David Briggs about.me/david_briggs http://about.me/david_briggs On 13 August 2014 12:40, Careina Edgooms 02531c126adf-dmarc-requ...@jiscmail.ac.uk wrote: Sorry for off topic question, just wondering if anyone has come across a study that shows the residue pka

Re: [ccp4bb] citation for shelxc and hkl2000

HKL2000 http://lmgtfy.com/?q=hkl2000+referencel=1 ShelxC http://lmgtfy.com/?q=ShelxC+referencel=1 HTH, Dave [image: David Briggs on about.me] David Briggs about.me/david_briggs http://about.me/david_briggs On 5 August 2014 09:47, Faisal Tarique faisaltari...@gmail.com wrote: Dear all

Re: [ccp4bb] ITC with heterogeneous protein

Hi Sajid, *Assuming* you have one site per monomer (rather than, say, one site per dimer), and *assuming* each binding event is completely independent ( I.e no co-operativity), you might just get away with running the experiment with the heterogeneous material. However, you might not be able to

Re: [ccp4bb] Salt!

? Failing that - try altering the construct - add / remove a few amino acids at each end, truncate any predicted loops... HTH, Dave [image: David Briggs on about.me] David Briggs about.me/david_briggs http://about.me/david_briggs On 16 July 2014 15:55, Bishop, Catherine E. cati...@ou.edu wrote

Re: [ccp4bb] How to remove phospholipids bound to a cytoplasmic protein

Hi Lionel, A few musings/suggestions If they are bound *inside* the protein, this suggests that the phospholipids might be very tightly bound. Do you have an affinity tag on your protein (e.g. His tag)? Perhaps you might immobilise the protein on a suitable resin and wash with copious amounts

Re: [ccp4bb] Www

Solubility issues though? Dr David C Briggs PhD http://about.me/david_briggs On 13 Jun 2014 11:45, Ian Clifton ian.clif...@chem.ox.ac.uk wrote: avinash singh avns.si...@gmail.com writes: wwdwwwy Wow, imagine the fluorescence from that… -- Ian ◎

Re: [ccp4bb] PDB passes 100,000 structure milestone

So, a bit like Fold-it but with actual data? :-D Dr David C Briggs PhD http://about.me/david_briggs On 16 May 2014 06:19, Pavel Afonine pafon...@gmail.com wrote: What about structures that are obviously wrong based on inspection of the density, but no one has bothered to challenge yet? The

Re: [ccp4bb] metal ion coordination

Hi Faisal, Take a look at Marjorie Harding's website http://tanna.bch.ed.ac.uk Loads of information there. Hth, Dave Dr David C Briggs PhD http://about.me/david_briggs On 17 Apr 2014 21:16, Faisal Tarique faisaltari...@gmail.com wrote: Dear all Can anybody please explain what is the

Re: [ccp4bb] Help in getting source of HOLE programme

AFAIK, Dr Smart now works at Global phasing. https://www.globalphasing.com/people/osmart/ HTH, Dave Dr David C Briggs PhD http://about.me/david_briggs On 12 Mar 2014 19:46, Appu kumar appu.kum...@gmail.com wrote: Hello everyone, I request all of you to please

Re: [ccp4bb] Large Conformational Change Upon Binding Ligand...

Integrin-ligand binding? See for example: http://humphrieslab.info/integrins.html Dr David C Briggs PhD http://about.me/david_briggs On 27 Feb 2014 19:43, Keller, Jacob kell...@janelia.hhmi.org wrote: Dear Crystallographers, Does anyone know of good examples of large, reversible

Re: [ccp4bb] Protein behaves as dimers in 2M NaCl and as high oligomers with 300mM NaCl

Hi Tom, Are your protein or related proteins known to dimerise? Is the dimer perhaps the natural state, and are the high oligomers non-specific aggregation? That would be my first guess, knowing nothing about the protein you are working on. If you are absolutely certain that the protein should

Re: [ccp4bb] KD of dimerization, off topic

Hi Careina, I'm not sure you can assume that the ratio of monomer and dimer will stay constant through the column - as you say, the protein is diluted during the run, the ratio will change, unless you have a super tight dimer - which clearly you do not. Also, as the mass and the molar extinction

[ccp4bb] Isolation of protein-protein complexes.

Dear all, sorry for the slightly off topic post, I have 2 proteins that have been shown to interact, by multiple groups, and by multiple techniques - namely ELISA, SPR and DPI. The Kd of the interaction as determined by SPR is on the order of 1 nM. I would very much like to crystallise this

Re: [ccp4bb] Isolation of protein-protein complexes.

interactions with the blank surface were seen Thanks once again everyone, Dave On 21 Jan 2014 15:51, David Briggs drdavidcbri...@gmail.com wrote: Dear all, sorry for the slightly off topic post, I have 2 proteins that have been shown to interact, by multiple groups, and by multiple techniques

Re: [ccp4bb] Best compounds for heavy atom soaks

Hi Rhys, It's worth paying close attention to your crystallisation conditions as well - some heavy atom compounds will not be at all soluble in very alkaline (they'll form insoluble hydroxides) or phosphate/sulphate containing mother liquors. A very low pH may reduce the binding efficiency of

Re: [ccp4bb] largest protein crystal ever grown?

Following on from John's comment, when I did my PhD at Birkbeck in the early 2000s, one of David Moss's other PhD students (John Bond) grew some gigantic (1cm edges) crystals of things like HEWL Myoglobin, which he then (somewhat perversely) crushed to load into capillaries for powder diffraction

Re: [ccp4bb] Structural transmission of signal across a membrane

Integrins? Loads of structures and models . On 24 Sep 2013 16:49, Edward A. Berry ber...@upstate.edu wrote: Gloria Borgstahl wrote: Hello ccp4ers, I am helping a colleague develop a grant and have a vague recollection of structures of transmembrane protein receptors that signal across the

Re: [ccp4bb] popular piece on X-ray crystallography

Following on from that - readers may be interested in Stephen Curry's post in the Guardian, regarding the Crystallography exhibit at the London Science Museum. http://www.guardian.co.uk/science/occams-corner/2013/apr/19/1 regards, Dave David C. Briggs PhD

Re: [ccp4bb] PDB crawler

Hi Sebastiano, Phyre Alarm would do something similar to what you suggest. http://www.sbg.bio.ic.ac.uk/phyre2/html/help.cgi?id=help/phyrealarm HTH, Dave David C. Briggs PhD http://about.me/david_briggs On 20 March 2013 12:40, Sebastiano Pasqualato

Re: [ccp4bb] Expert opinion for optimizing ethylene glycol crystallization condition

Hi Shankar, I don't claim to be an expert, but I've used ethylene glycol as a cryoprotectant many times, and 30% (v/v) would probably be sufficient for proper freezing without any further addition. Did you try and shoot a crystal at room temperature? Hth, Dave On Jan 2, 2013 5:59 PM,

Re: [ccp4bb] Izit dye stained crystal

Hi Sarathy, Are you sure your anomalous scatterers are not the Ca in the crystallisation buffer? These would also bind to the acidic residues in your protein, and Ca has a greater f'' (~2.5e compared to less than 1.5e for S) at the wavelength you used. Just another possibility - unless you

Re: [ccp4bb] ResearchGate?

Hello Adrian, I use Research Gate and there are a few occasions where I have found it useful, particularly the questions feature. HTH, Dave David C. Briggs PhD http://about.me/david_briggs On 30 October 2012 16:13, Adrian Goldman adrian.gold...@helsinki.fi wrote:

Re: [ccp4bb] Mercury phenylglyoxal

Hi Vijay, Not an answer to your question, but I tried and failed to source any about 10 years ago. I had a chemist friend try to make some following the a protocol we found on the web - which didn't work. The consensus on the ccp4bb back then was that it was a bit of mythical beast, and I never

Re: [ccp4bb] Do my SAXS data agree with the crystal structure?

Dear Xun, Regarding your monomer vs dimer, theoretical vs observed crysol plots - yes - they are significantly different. If you focus at the very lowest q part of the curve - the deviation there in your monomer plots indicate that there is a significant size difference between your PX monomer

Re: [ccp4bb] Do my SAXS data agree with the crystal structure?

Hi Xun, it is difficult to judge without seeing the P(r) plots, but seeing as you have a dimer in your crystal structure and a dimer in your SAXS, AND your Chi2 value seems reasonable for a good match between PX and SAXS, I'd say you've got what you need. A matching P(r) plot would be nice, but

Re: [ccp4bb] very informative - Trends in Data Fabrication

Trollus, Trollum, Trolli, Trollo, Trolli, Trollos, Trollorum, Trollis. David C. Briggs PhD Father, Structural Biologist and Sceptic University of Manchester E-mail: david.c.bri...@manchester.ac.uk Webs :

Re: [ccp4bb] Unable to reproduce robot tray hits in hand trays

Hi Matt, This doesn't really answer your question directly, but sidesteps around the issue - I wrote a little something on this exact subject not so long ago - http://xtaldave.wordpress.com/2012/02/23/on-protein-crystallisation/ (You can ignore the first 5 paragraphs of intro - it was written

[ccp4bb] Envelope Phasing.

Hi CCP4bb, I would like to ask about envelope phasing - specifically with SAXS data. There are papers (1) and tutorials (2) describing how this might be done, but I have also found comments on the ccp4bb, such as this one (http://www.proteincrystallography.org/ccp4bb/message11690.html) which are

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