[ccp4bb] cc minor off topic protein symmetry

Many thanks to Mark van Raaij for a super example - 5EFV - clear heterogeneity at the hinge/kink; although this then doesn't return to the 1st axis Its interesting to contemplate proteins doing this several times in a physiologically "real" way (the models I'm looking at do this) - they need gly

[ccp4bb] cc - minor off topic protein symmetry

(I should note that my use of symmetry is used on the protein level rather on the absolute crystallographic equivalence level - imagine this trimer sat within a P1 cell, and that minor wobbles generate pseudosymmetry in places anyway - what I'm interested in is the topology of the chain at the j

[ccp4bb] minor off topic protein symmetry

Dear wise list, I have a question regarding protein oligomers that have multiple, differing axes of symmetry - stimulated by some perplexing but likely real Alphafold models. I think it's the protein equivalent of this old chestnut: https://en.wikipedia.org/wiki/Three_utilities_problem Consid

[ccp4bb] Re [ccp4bb] Crystallizing a tough target

Hi Kavya A few things I would try: -set up your plates (and incubate) at 4 degrees - perhaps in a cold room -use Alphafold model to identify patches of charge that you can mutate -perhaps even introduce a few sticky motifs (e.g. herein and ref 26 https://www.ncbi.nlm.nih.gov/pmc/articles/PMC2567

[ccp4bb] PDBeMotif and like methods

Dear CCP4 list, I have just tried accessing the "old" EBI PDBeMotif link and it is non-functional. Assuming that it may or may not be moved/unavailable, I am asking what your favourite methods are for mapping similar residue interactions in dissimilar folds i.e. if you have a nice network of 3

[ccp4bb] Off topic: docking for biological insight

Dear ccp4 community, When I see the (improving) accuracy of docking in generating drug like leads, I see predominantly synthetic compounds used. It occurred to me that it might be informative to use docking of naturally occurring compounds (amino acids, sugars, lipids, factors etc) to find pocke

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

Hi Scott We have obtained a structure of a flexible clamshell like fold only after using a disulphide mutant to lock the domains based on a Rosetta Fold model. Interestingly, Alphafold put the very same residues further apart (probably a relevant "open" pose) Andy ##

[ccp4bb] Off topic - gene toxic for expression strains

Dear Board, Perhaps off-topic, but in the wider scope it's relevant to many on here. We have a gene that we are able to clone, and propagate in DH5a etc non-expression cells (hence nucleotide sequence is non-toxic) But, when we attempt to transfer to an expression strain we get no colonies We

[ccp4bb] UV Microscope for Crystal Imaging

Dear All Does anyone have a recommendation for a joint light/UV microscope for manual Crystal screen inspection (i.e. a stand alone, not linked to automatation etc)? Positive (ccp4bb) and negative (direct) impressions welcome! Thanks Andy ###

Re: [ccp4bb] self-inhibitory fragments from a protein - clarification

Dear All just to clarify, I'd be most interested in examples resulting from proteolysis, rather than say, packing effects Best Andy To unsubscribe from the CCP4BB list, click the following link: https://www.jiscmail.ac.uk

[ccp4bb] self-inhibitory fragments from a protein

Dear CCP4bb, I have a partly specific question, relating to multidomain enzymes. We have a nice hi-res structure of an enzyme, which comprises domain A (non-catalytic) and domain B (enzymatic) Through the magic of unstimulated in-tray proteolysis, there are one and a half copies in the ASU, on

[ccp4bb] Elemental Analysis

Dear CCP4ers We have a protein with moderate resolution (mid 2s) with obvious metal ion density holding the oligomer together. We can make an educated guess that the ions are likely to be Mg/Ca and can look at co-ordination and bond lengths.but I have a more specific question - would it be

Re: [ccp4bb] Best protocols to advance a low resolution twin

To follow on from this thread: To answer Jon, I did try to see if P6 subgroups were a possibility but can rule this out for a few reasons (MR doesn't give solutions, merging stats not suggestive of P6, the other dataset with the twin fraction that is significantly further from 50:50); and the d

[ccp4bb] Best protocols to advance a low resolution twinned MR map

Dear all, I've a project with two historical datasets (i.e. not sure of when/if I can reproduce to solve problem by getting better data) that are twinned. The spacegroup is P32, with operator -h, -k, l cell = 83.5 83.5 64.2 90 90 120 protein has three domains d1-d2-d3, likely 50% solvent, 2

[ccp4bb] Thanks and outcome of ([ccp4bb] Looking for method to find similar oligomeric arrangement)

Dear All Many thanks to Johann for the suggestion of Topsearch - rather pleasingly our tetramer is novel in the tetrameric arrangement, but each of the dimers has its DNA recognition helices in agreement with several hits 😊a win all round Bringing joy to my lockdown. Best Andy ###

Re: [ccp4bb] Looking for method to find similar oligomeric arrangement

Thanks to all of you pointing me towards ProtCID, but unless I'm mistaken this works only for deposited co-ordinates / PDB entries? I was more looking for something that would take my (yet to be deposited) PDB file, with its inherent tetrametric hth fold, and find similar folds in similar arra

Re: [ccp4bb] Looking for method to find similar oligomeric arrangement

Thanks Boaz for the SSM heads up. I was much too DALI-centric! One last point, if anyone can offer an opinion - I presume most methods will match but only within ASU contents. Does any approach utilize deposited "biological unit" classification or (better still) crystallographic symmetry oper

[ccp4bb] Looking for method to find similar oligomeric arrangement

Dear colleagues, I have a structure of a simple, common fold (hth, dna-binding) that I believe has oligomerized in a different way to that observed for any members of the superfamily that I can reasonably analyze. So if I run fold comparison analysis (e.g. DALI) it will find similar structur

[ccp4bb] RE [ccp4bb] Intergrown crystals and excess of nucleation

Hi Nikolas The first thing I would try is to solubilise the protein a bit more, maybe by adding a few % (1-5%) glycerol or ethylene glycol to the growth conditions when you set up the drops. The next thing would be to experiment with ratio (protein:precipitant) and drop size. Lastly, trial ad

[ccp4bb] off topic: Membrane protein "into" soluble protein

Dear ccp4bb members, A quick off topic question. I have a protein whose domain structure runs N-TM-receptor-TM-enzyme-C, i.e. is a two transmembrane-helix containing signalling protein. One would suspect that the receptor domain has ends that cluster, and that the enzyme (via homology with know

[ccp4bb] multi-xtal averaging

Dear all, Thanks to Vincent, Eleanor, herman and others! It turns out I've managed to sort it out a different way - the initial TFZ of 10 with cutout density and resultant map was a bit underwhelming and indicative of an issue - so I cut my ~140ang long molecule in half and searched with densit

[ccp4bb] FW: complex multi-crystal averaging

Ok - small oversight - I can see now that phenix has the atoms placed/saved during cutout stage - so how best to use PHASER output to shift this file? Andy From: Andrew Lovering (School of Biosciences) Sent: 10 January 2019 10:48 To: 'ccp4bb@jiscmail.ac.uk' Subject: RE: complex mul

Re: [ccp4bb] complex multi-crystal averaging

xtal1 to match placed density of MR in xtal2" :) Andy From: Andrew Lovering (School of Biosciences) Sent: 09 January 2019 16:08 To: 'ccp4bb@jiscmail.ac.uk' Subject: complex multi-crystal averaging Dear All, I suspect the way out of this is a new crystal (!) but interested to h

[ccp4bb] complex multi-crystal averaging

Dear All, I suspect the way out of this is a new crystal (!) but interested to hear any advice. I have two crystal forms of a 500aa protein, vaguely tube-shaped 1=P3121, diffracts to 4.1 ang, 3 copies in asu, 86% solvent; map indicates it is a relative of other folds (but those are not so clos

[ccp4bb] Postdoctoral Position Available

Dear All, A Wellcome-Trust postdoctoral position (for 2 years, possibly extendible to 4) is open in the Lovering laboratory, using x-ray crystallography to study the means by which predatory bacteria can form a pore in the prey outer membrane. The link for the position is https://bit.ly/2EET1ta

Re: [ccp4bb] coiled-coil "degree"

Dear Jorge This may not be exactly what you require, but there is a nice example of analysis in the shift of a coiled coil register herein: Elife. 2018 Jun 5;7. pii: e34815. doi: 10.7554/eLife.34815. Asymmetric activation mechanism of a homodimeric red light-regulated photoreceptor. Best, Andy

[ccp4bb] Mild Denaturation to lose ligand

Dear All, I have a protein from which I'd like to lose the co-purifying ligand (Coenzyme A). The protein is his-tagged. The plan here is to bind it to a resin and then run over a mild denaturant solution to encourage ligand loss (it seems that extended dialysis does not achieve this). Our star

Re: [ccp4bb] blast pdb for a very short sequence (3 amino acids)

The backbone angle search in pdbemotif should limit it to particular secondary structure. Good luck! Andy From: xiaolei...@gmail.com [xiaolei...@gmail.com] Sent: October 26, 2017 5:53 PM To: Andrew Lovering; Joana Pereira Cc: CCP4BB@JISCMAIL.AC.UK Subject

Re: [ccp4bb] blast pdb for a very short sequence (3 amino acids)

I would guess that you want to search PDB for a small motif you've observed in a structure of yours, looking to find examples of it used in same context? If that's true, I would recommend PDBemotif, put the sequence in, and then use the backbone psi phi angles of your motif as a secondary constra

Re: [ccp4bb] unusual monoclinic relation?

] Sent: 19 December 2016 19:58 To: Andrew Lovering Cc: CCP4BB@jiscmail.ac.uk Subject: Re: [ccp4bb] unusual monoclinic relation? Dear Andrew, did you remove all cysteins, and all methionines with the mutations? Your resolution is about 2A, if I understand correctly. This may be suitable for S

Re: [ccp4bb] unusual monoclinic relation?

e for 2017! p.s. no anomalous signal in there - ironically mutant we're using knocks bound metal out! Thanks again Andy From: herman.schreu...@sanofi.com [mailto:herman.schreu...@sanofi.com] Sent: 19 December 2016 16:26 To: Andrew Lovering; CCP4BB@JISCMAIL.AC.UK Subject: AW: unusual monoclini

Re: [ccp4bb] unusual monoclinic relation?

night and so I killed the job thinking it was thrashing Andy From: herman.schreu...@sanofi.com [mailto:herman.schreu...@sanofi.com] Sent: 19 December 2016 10:43 To: Andrew Lovering; CCP4BB@JISCMAIL.AC.UK Subject: AW: unusual monoclinic relation? Dear Andrew, Just a few questions: -Do the proce

[ccp4bb] unusual monoclinic relation?

Dear All, I have just collected data on a mutant of a protein that should be facile to solve by molrep (one residue/320 changed, approx 2Ang resolution) but is proving problematic. Data merging stats look good. The spacegroup is monoclinic, P21, the cell: a=39.47 b=157.36 c=74.9 beta=98.26 I

[ccp4bb] Postdoctoral Position, University of Birmingham UK

(posted on behalf of Dr Anne-Marie Krachler) Dear Colleagues, Description for a postdoctoral position in the lab of Dr Anne-Marie Krachler, University of Birmingham, UK; Post is available from 1 March 2014 Postdoctoral Research Fellow (Structural Biology) Applications are invited for a highly mot

[ccp4bb] Postdoctoral Position at Birmingham, UK

Dear Colleagues, Description for a postdoctoral position in the lab of Andrew Lovering, University of Birmingham, UK; work following on from our recent publication in Plos Pathogens: http://www.plospathogens.org/article/info%3Adoi%2F10.1371%2Fjournal.ppat.1002524 Funded by the BBSRC and in