Re: [ccp4bb] issues
Dear Charles, Since my institution migrated from lotus notes to MS Outlook many (not all) ccp4bb mails get into quarantine and I have to liberate them manually from the list. This might be the reason why ccp4bb black-flagged my e-mail address this morning (first time since almost 30 years). Of course, I put the ccp4bb e-mail address and the Jismail domain on the list of allowed senders, but this doesn't seem to work properly. It seems to reduce the number of quarantine events, but not all. According to my local IT guys and the guys one level above there is no way to solve this issue locally. Perhaps CCP4BB could talk to some Microsoft officials, trying to convince them that we are all good guys (most likely ;-). -Peer According to my local IT guys and the guys one level above there is only one way to so On 10.01.2023 10:49, Charles Ballard - STFC UKRI wrote: Dear All Jisc, who we use to run the ccp4bb have been having issues delivering to some (a lot) of e-mail addresses. So far 1400 addresses have been unsubscribed, and another 2000 are being monitored. I have turned for automatic removal (I think) and asked that the status of the list be returned to that Saturday’s members lists. All the best Charles This email and any attachments are intended solely for the use of the named recipients. If you are not the intended recipient you must not use, disclose, copy or distribute this email or any of its attachments and should notify the sender immediately and delete this email from your system. UK Research and Innovation (UKRI) has taken every reasonable precaution to minimise risk of this email or any attachments containing viruses or malware but the recipient should carry out its own virus and malware checks before opening the attachments. UKRI does not accept any liability for any losses or damages which the recipient may sustain due to presence of any viruses. To unsubscribe from the CCP4BB list, click the following link: https://www.jiscmail.ac.uk/cgi-bin/WA-JISC.exe?SUBED1=CCP4BB=1 <https://www.jiscmail.ac.uk/cgi-bin/WA-JISC.exe?SUBED1=CCP4BB=1> -- - PD Dr. Peer Mittl Department of Biochemistry Room 44M03 University Zürich Winterthurerstr. 190 8057 Zürich Switzerland Phone: +41-44-635 6559 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] ISO model with large groups of atoms in alternative conformations
Dear Pavel, Recently we determined the structure of a fusion protein with 2.5 chains in the AU in P3(2)21 (I posted this case in the ccp4bb). The protein forms a pentamer with one chain sitting on the 2-fold. I refined it in P3(2) with 4 well defined chains and the 5th chain having two conformations. This 5th chain interacts exclusively with the other 4 chains of the pentamer, rendering it asymmetric, but this asymmetry is not transmitted to the crystal contacts of the pentamer. My interpretation is that the pentamer packs in a P3(1)21 lattice with a random distribution of the 5th chain. We tried many other SGs (including P1) with and without twinning, but it always came back to this interpretation. All the best, Peer Am 24.08.2022 um 00:02 schrieb Pavel Afonine: Dear community, I’m looking for an example of a crystal structure where a large group of atoms (as large as a whole chain or even a domain) have more than one distinct conformation that would require modeling of such chain/domain as more than one individual copy, with each copy having partial occupancy. I’m not sure if that even exists but if someone can share an example that'd be very much appreciated! Thanks! Pavel To unsubscribe from the CCP4BB list, click the following link: https://www.jiscmail.ac.uk/cgi-bin/WA-JISC.exe?SUBED1=CCP4BB=1 <https://www.jiscmail.ac.uk/cgi-bin/WA-JISC.exe?SUBED1=CCP4BB=1> -- ** PD Dr. Peer Mittl Biochemisches Institut Universität Zürich 8057 Zürich Switzerland Phone: +41-44-635 6559 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] Antwort: [ccp4bb] AW: [ccp4bb] AW: [ccp4bb] Antwort: Re: [ccp4bb] chain on 2-fold axis?
ed chain are randomly distributed throughout the crystal. Given that the two orientations have different twin fractions, my bet is that the twin supporters are right. Best, Herman Von: Lijun Liu mailto:lijunli...@gmail.com>> Gesendet: Freitag, 27. August 2021 15:57 An: Schreuder, Herman /DE <mailto:herman.schreu...@sanofi.com>> Cc: ccp4bb@jiscmail.ac.uk <mailto:ccp4bb@jiscmail.ac.uk> Betreff: Re: [ccp4bb] AW: [ccp4bb] Antwort: Re: [ccp4bb] chain on 2-fold axis? Dear Herman: if you say “twinned” chains, then it already means same thing of the two and the side chain interactions could not be different (as you can see only one copy in the output coordinates), unless you talking about borders between twin pieces. In this situation, the only I could imagine to be if P3221, it would have to make that chain very very rapidly jump between two states in the same asu, which can be easily proven wrong. I do agree if refined under P3221 without restraining the two strictly, side chain interactions may show differences —— data always not perfect! Lijun Sent from my iPhone On Aug 27, 2021, at 8:12 AM, Schreuder, Herman /DE mailto:herman.schreu...@sanofi.com>> wrote: Dear Lijun, with this argument I agree: the interactions between the two orientations of the “twinned” chain and the neighboring molecules will be different and the interacting side chains will almost certainly have different orientations, which necessitates a twinning of the whole structure. Best, Herman Von: CCP4 bulletin board <mailto:CCP4BB@JISCMAIL.AC.UK>> Im Auftrag von Lijun Liu Gesendet: Freitag, 27. August 2021 14:22 An: CCP4BB@JISCMAIL.AC.UK <mailto:CCP4BB@JISCMAIL.AC.UK> Betreff: Re: [ccp4bb] AW: [ccp4bb] Antwort: Re: [ccp4bb] chain on 2-fold axis? I believe it is a twin from P32. Not like the assignment of double conformations with partial occupancies to small part of asu, for examples, a side chain of lysin or a small fragment of a protein, which have both conformations stayed in the same specific asu at the same time. For this p3221 asu, if one copy of that chain occupies the part of the asu, the other so-called conformation will not appear in the same asu at the same time, so the conformation and occupancy issue was arisen from different cell units, it is a twin from p32. Talking about the asu of p3221, it holds 3 chains, the chain of interest assigned as 2 conformations related by a strict 2-fold. Since you believe and have reduced to P3221, then you should restrain the two overall strictly (since absolutely most of these atoms are not on special positions) and not to refine the two 0.5 occupancies either, respecting the crystallographic 2-fold. This is equivalent to perfect twin from p32 theoretically (although refinement with twin mode in p32 and none-twin mode in p3221 may give difference even large). If you refined and resulted in occupancies away from 0.5, the symmetry was proven to be broken, which supports P32 twinning. Lijun Sent from my iPhone On Aug 27, 2021, at 6:56 AM, Oganesyan, Vaheh <mailto:vaheh.oganes...@astrazeneca.com>> wrote: How P3221 can be an option if it assumes chain on axis? I guess I’m missing something, but per my belief only those sg will be possible for which there is no axis going through the extra molecule. P1 sg looks the only correct option here in my humble opinion. Democracy (voting) depends on science. However, the reverse is not, thankfully. Vaheh From: CCP4 bulletin board <mailto:CCP4BB@JISCMAIL.AC.UK>> On Behalf Of Peer Mittl Sent: Friday, August 27, 2021 6:32 AM To: CCP4BB@JISCMAIL.AC.UK <mailto:CCP4BB@JISCMAIL.AC.UK> Subject: Re: [ccp4bb] AW: [ccp4bb] Antwort: Re: [ccp4bb] chain on 2-fold axis? Dear Herman, The answer probably depends on the impact of the "extra" chain on the sublattice. If there is no impact the "true" space group is P3221 with one chain on the special position. If the swapping of the extra chain influences the sublattice P32 (or C2 or P1, as pointed out by Kay) twinned to P3221 might be the better description. All the best, Peer On 27.08.2021 10:56, Schreuder, Herman /DE wrote: > > Dear Peer and Eleanor, > > This is indeed what I am suspecting: If the “twinning operator” in P32 > puts 4 out of 5 protein chains on top of symmetry mates, is the “true” > space group then P32, with 5 twinned chains, or P3221 with 4 normal > chains and 1 chain on a special position? I would vote for the latter. > > Best, > > Herman > > *Von:* CCP4 bulletin board <mailto:CCP4BB@JISCMAIL.AC.UK>> *Im Auftrag von > *Peer Mittl > *Gesendet:* Freitag, 27. August 2021 10:17 > *An:* CCP4BB@JISCMAIL.AC.UK <mailto:CCP4BB@JISCMAIL.AC.UK> > *Betreff:* Re: [ccp4bb] Antwort: Re: [ccp4bb] chain on 2-fold axis? > > Dear Eleanor, > > I indeed used
[ccp4bb] Antwort: Re: [ccp4bb] AW: [ccp4bb] Antwort: Re: [ccp4bb] chain on 2-fold axis?
Dear Vaheh, I agree with you, at least in your last statement. I guess we all agree that certain molecules can occupy special positions on true rotation axis. Strictly, this is only possible if the molecule obeys the rotation symmetry. For water molecules on 2-folds you already have to make assumptions about the "invisible" protons. I guess, many of us have seen even larger and asymmetric solvent molecules, such as glycerol, MPD or buffer molecules on special positions, which locally break the crystal symmetry. The work around for this issue would be to define two alternative conformations for this molecule, because these conformations do not "see" each other, as pointed out by Herman. But where is the upper limit for the molecular weight of this molecule? I (and perhaps most other crystallographers) would not refine such a case as a twinned structure in a lower symmetry space group, because the major part of the AU obeys the crystal symmetry. Furthermore, there is a fundamental difference between crystallographic symmetry and the symmetry of a twin law. The crystallographic symmetry covers the entire crystal, whereas the twin law just relates twin domains locally. Refining a true P3221 structure as a twinned P32 structure is simply the wrong thing to do. All the best, Peer -"CCP4 bulletin board" schrieb: - An: CCP4BB@JISCMAIL.AC.UK Von: "Oganesyan, Vaheh" Gesendet von: "CCP4 bulletin board" Datum: 27.08.2021 13:56 Betreff: Re: [ccp4bb] AW: [ccp4bb] Antwort: Re: [ccp4bb] chain on 2-fold axis? How P3221 can be an option if it assumes chain on axis? I guess I’m missing something, but per my belief only those sg will be possible for which there is no axis going through the extra molecule. P1 sg looks the only correct option here in my humble opinion. Democracy (voting) depends on science. However, the reverse is not, thankfully. Vaheh From: CCP4 bulletin board On Behalf Of Peer Mittl Sent: Friday, August 27, 2021 6:32 AM To: CCP4BB@JISCMAIL.AC.UK Subject: Re: [ccp4bb] AW: [ccp4bb] Antwort: Re: [ccp4bb] chain on 2-fold axis? Dear Herman, The answer probably depends on the impact of the "extra" chain on the sublattice. If there is no impact the "true" space group is P3221 with one chain on the special position. If the swapping of the extra chain influences the sublattice P32 (or C2 or P1, as pointed out by Kay) twinned to P3221 might be the better description. All the best, Peer On 27.08.2021 10:56, Schreuder, Herman /DE wrote: > > Dear Peer and Eleanor, > > This is indeed what I am suspecting: If the “twinning operator” in P32 > puts 4 out of 5 protein chains on top of symmetry mates, is the “true” > space group then P32, with 5 twinned chains, or P3221 with 4 normal > chains and 1 chain on a special position? I would vote for the latter. > > Best, > > Herman > > *Von:* CCP4 bulletin board *Im Auftrag von > *Peer Mittl > *Gesendet:* Freitag, 27. August 2021 10:17 > *An:* CCP4BB@JISCMAIL.AC.UK > *Betreff:* Re: [ccp4bb] Antwort: Re: [ccp4bb] chain on 2-fold axis? > > Dear Eleanor, > > I indeed used r/tefmac for the refinement and it came up with the values > HKL (a=0.56), KH-L (a=0.44). It would be interesting to see if a > refinement in P3221 would come up with the same occupancies for the > alternative conformations for the "extra" chain on the 2-fold axis. It > seems as if the "well-ordered" chains (2 in P3221, 4 in P32) form a > sublattice with P3221 symmetry and it's just the "extra" chain, which > generates the twinning. > > All the best, > Peer > > On 26.08.2021 18:09, Eleanor Dodson wrote: > > Motto =mitti in predictive text! > > > > On Thu, 26 Aug 2021 at 16:52, Eleanor Dodson > > mailto:eleanor.dod...@york.ac.uk > <mailto:eleanor.dod...@york.ac.uk%20%3cmailto:eleanor.dod...@york.ac.uk>>> > wrote: > > > > Great, motto. I think you have nailed it! Did you use tefmac for > > twinned refinement? And if so what did it suggest the twin > > fraction is? > > > > On Thu, 26 Aug 2021 at 16:30, Peer Mittl <mailto:mi...@bioc.uzh.ch%0b>> <mailto:mi...@bioc.uzh.ch > <mailto:mi...@bioc.uzh.ch>>> wrote: > > > > Yes, the data indeed seems to be twinned and the tNCS has > > masked the twinning statistics, which is why I haven't > > considered it so far. > > > > I have not tried twinned refinement in C2 and P1 yet, but > > refining 4 chains in P32 with twinning yields a difference ED > > map that clearly indicates one (and just on!) orientation for > > the 5th chain. Thank you all for your suggestions. > > &
Re: [ccp4bb] AW: [ccp4bb] Antwort: Re: [ccp4bb] chain on 2-fold axis?
Dear Herman, The answer probably depends on the impact of the "extra" chain on the sublattice. If there is no impact the "true" space group is P3221 with one chain on the special position. If the swapping of the extra chain influences the sublattice P32 (or C2 or P1, as pointed out by Kay) twinned to P3221 might be the better description. All the best, Peer On 27.08.2021 10:56, Schreuder, Herman /DE wrote: Dear Peer and Eleanor, This is indeed what I am suspecting: If the “twinning operator” in P32 puts 4 out of 5 protein chains on top of symmetry mates, is the “true” space group then P32, with 5 twinned chains, or P3221 with 4 normal chains and 1 chain on a special position? I would vote for the latter. Best, Herman *Von:* CCP4 bulletin board *Im Auftrag von *Peer Mittl *Gesendet:* Freitag, 27. August 2021 10:17 *An:* CCP4BB@JISCMAIL.AC.UK *Betreff:* Re: [ccp4bb] Antwort: Re: [ccp4bb] chain on 2-fold axis? Dear Eleanor, I indeed used r/tefmac for the refinement and it came up with the values HKL (a=0.56), KH-L (a=0.44). It would be interesting to see if a refinement in P3221 would come up with the same occupancies for the alternative conformations for the "extra" chain on the 2-fold axis. It seems as if the "well-ordered" chains (2 in P3221, 4 in P32) form a sublattice with P3221 symmetry and it's just the "extra" chain, which generates the twinning. All the best, Peer On 26.08.2021 18:09, Eleanor Dodson wrote: > Motto =mitti in predictive text! > > On Thu, 26 Aug 2021 at 16:52, Eleanor Dodson > mailto:eleanor.dod...@york.ac.uk <mailto:eleanor.dod...@york.ac.uk%20%3cmailto:eleanor.dod...@york.ac.uk>>> wrote: > > Great, motto. I think you have nailed it! Did you use tefmac for > twinned refinement? And if so what did it suggest the twin > fraction is? > > On Thu, 26 Aug 2021 at 16:30, Peer Mittl <mailto:mi...@bioc.uzh.ch%0b>> <mailto:mi...@bioc.uzh.ch <mailto:mi...@bioc.uzh.ch>>> wrote: > > Yes, the data indeed seems to be twinned and the tNCS has > masked the twinning statistics, which is why I haven't > considered it so far. > > I have not tried twinned refinement in C2 and P1 yet, but > refining 4 chains in P32 with twinning yields a difference ED > map that clearly indicates one (and just on!) orientation for > the 5th chain. Thank you all for your suggestions. > > Have a nice evening, > Peer > > -"CCP4 bulletin board" <mailto:CCP4BB@JISCMAIL.AC.UK%0b>> <mailto:CCP4BB@JISCMAIL.AC.UK <mailto:CCP4BB@JISCMAIL.AC.UK>>> schrieb: - > An: CCP4BB@JISCMAIL.AC.UK <mailto:CCP4BB@JISCMAIL.AC.UK> <mailto:CCP4BB@JISCMAIL.AC.UK <mailto:CCP4BB@JISCMAIL.AC.UK>> > Von: "Kay Diederichs" > Gesendet von: "CCP4 bulletin board" > Datum: 26.08.2021 16:41 > Betreff: Re: [ccp4bb] chain on 2-fold axis? > > Dear Peer, > > I suspect that the true spacegroup has lower symmetry than > P3221, and that there may be twinning masked by tNCS. > Subgroups of P3221 are C2 and P32 ( > https://strucbio.biologie.unikonstanz.de/xdswiki/index.php/Space_group_determination#Subgroup_and_supergroup_relations_of_these_space_groups <https://strucbio.biologie.unikonstanz.de/xdswiki/index.php/Space_group_determination#Subgroup_and_supergroup_relations_of_these_space_groups> > <https://strucbio.biologie.unikonstanz.de/xdswiki/index.php/Space_group_determination#Subgroup_and_supergroup_relations_of_these_space_groups <https://strucbio.biologie.unikonstanz.de/xdswiki/index.php/Space_group_determination#Subgroup_and_supergroup_relations_of_these_space_groups>> > ) > and of course P1. > What I'd do is process the data, and solve (use the best chain > of the refined P3221 model for MR) and refine the structure in > these spacegroups. > Inspect the results: If P1 is clearly better than P32 and C2, > P1 is correct. > If C2 (P32) is clearly better than P32 (C2), then P1 should > give the same R-values as the better one; if so, P1 can be > discarded. > Try this with and without twin refinement - although it's hard > to compare R-values of non-twinned and twinned refinements. > > The automatic way to do this is with Zanuda. If you run that > locally, you can make refmac do twin refinement. > > For all resulting structures, I'd also feed the resulting > Fcalc (!) into pointless. That should reveal that the packing > is indeed close to P3221. > > Best wishes, > Kay > > > On Thu, 26 Aug 2021 11:54:06 +0200, Peer Mittl > mailto:mi...@bioc.uzh.ch <mailto:mi...@bioc.uzh.ch%20%3cmailto:mi...@bioc.uzh.ch>>> wrote: > > >Der CCP4 community, > > > >Is there a refinement program that can handle protein > monomers sitting &g
Re: [ccp4bb] Antwort: Re: [ccp4bb] chain on 2-fold axis?
Dear Eleanor,
I indeed used r/tefmac for the refinement and it came up with the values
HKL (a=0.56), KH-L (a=0.44). It would be interesting to see if a
refinement in P3221 would come up with the same occupancies for the
alternative conformations for the "extra" chain on the 2-fold axis. It
seems as if the "well-ordered" chains (2 in P3221, 4 in P32) form a
sublattice with P3221 symmetry and it's just the "extra" chain, which
generates the twinning.
All the best,
Peer
On 26.08.2021 18:09, Eleanor Dodson wrote:
Motto =mitti in predictive text!
On Thu, 26 Aug 2021 at 16:52, Eleanor Dodson
mailto:eleanor.dod...@york.ac.uk>> wrote:
Great, motto. I think you have nailed it! Did you use tefmac for
twinned refinement? And if so what did it suggest the twin
fraction is?
On Thu, 26 Aug 2021 at 16:30, Peer Mittl mailto:mi...@bioc.uzh.ch>> wrote:
Yes, the data indeed seems to be twinned and the tNCS has
masked the twinning statistics, which is why I haven't
considered it so far.
I have not tried twinned refinement in C2 and P1 yet, but
refining 4 chains in P32 with twinning yields a difference ED
map that clearly indicates one (and just on!) orientation for
the 5th chain. Thank you all for your suggestions.
Have a nice evening,
Peer
-"CCP4 bulletin board" mailto:CCP4BB@JISCMAIL.AC.UK>> schrieb: -
An: CCP4BB@JISCMAIL.AC.UK <mailto:CCP4BB@JISCMAIL.AC.UK>
Von: "Kay Diederichs"
Gesendet von: "CCP4 bulletin board"
Datum: 26.08.2021 16:41
Betreff: Re: [ccp4bb] chain on 2-fold axis?
Dear Peer,
I suspect that the true spacegroup has lower symmetry than
P3221, and that there may be twinning masked by tNCS.
Subgroups of P3221 are C2 and P32 (
https://strucbio.biologie.unikonstanz.de/xdswiki/index.php/Space_group_determination#Subgroup_and_supergroup_relations_of_these_space_groups
<https://strucbio.biologie.unikonstanz.de/xdswiki/index.php/Space_group_determination#Subgroup_and_supergroup_relations_of_these_space_groups>
)
and of course P1.
What I'd do is process the data, and solve (use the best chain
of the refined P3221 model for MR) and refine the structure in
these spacegroups.
Inspect the results: If P1 is clearly better than P32 and C2,
P1 is correct.
If C2 (P32) is clearly better than P32 (C2), then P1 should
give the same R-values as the better one; if so, P1 can be
discarded.
Try this with and without twin refinement - although it's hard
to compare R-values of non-twinned and twinned refinements.
The automatic way to do this is with Zanuda. If you run that
locally, you can make refmac do twin refinement.
For all resulting structures, I'd also feed the resulting
Fcalc (!) into pointless. That should reveal that the packing
is indeed close to P3221.
Best wishes,
Kay
On Thu, 26 Aug 2021 11:54:06 +0200, Peer Mittl
mailto:mi...@bioc.uzh.ch>> wrote:
>Der CCP4 community,
>
>Is there a refinement program that can handle protein
monomers sitting
>on crystallographic 2-folds?
>
>This is probably a strange question but we have the following
situation.
>We have a 2.6 Ang datasets in SG P3221 (Rpim=4%, Isa=19.6)
and a clear
>molrep solution with 2 chains, albeit with tNCS (0/0/0.5)
that can be
>refined to around 27/33% Rfactor. According to Vm a third
chain could be
>present. So far so good, but there is clear difference ED for
a third
>chain sitting exactly on the 2-fold. Since the protein has a
peculiar
>shape, one can tell even its orientation. I can relax the
symmetry to
>P32 (or even P1) and place the missing chain with 50%
occupancy on the
>2-fold. This model can be refined, but I do not like this
work around,
>because the data is clearly P3221.
>
>Any hints on similar crystal pathologies and how they have
been handled
>would be helpful.
>
>All the best,
>Peer
>
>
>
>To unsubscribe from the CCP4BB list, click the following link:
>https://www.jiscmail.ac.uk/cgi-bin/WA-JISC.exe?SUBED1=CCP4BB=1
<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 <http://www.jiscmail.ac.uk/CCP4BB>,
[ccp4bb] Antwort: Re: [ccp4bb] chain on 2-fold axis?
Yes, the data indeed seems to be twinned and the tNCS has masked the twinning statistics, which is why I haven't considered it so far. I have not tried twinned refinement in C2 and P1 yet, but refining 4 chains in P32 with twinning yields a difference ED map that clearly indicates one (and just on!) orientation for the 5th chain. Thank you all for your suggestions. Have a nice evening, Peer -"CCP4 bulletin board" schrieb: - An: CCP4BB@JISCMAIL.AC.UK Von: "Kay Diederichs" Gesendet von: "CCP4 bulletin board" Datum: 26.08.2021 16:41 Betreff: Re: [ccp4bb] chain on 2-fold axis? Dear Peer, I suspect that the true spacegroup has lower symmetry than P3221, and that there may be twinning masked by tNCS. Subgroups of P3221 are C2 and P32 ( https://strucbio.biologie.unikonstanz.de/xdswiki/index.php/Space_group_determination#Subgroup_and_supergroup_relations_of_these_space_groups ) and of course P1. What I'd do is process the data, and solve (use the best chain of the refined P3221 model for MR) and refine the structure in these spacegroups. Inspect the results: If P1 is clearly better than P32 and C2, P1 is correct. If C2 (P32) is clearly better than P32 (C2), then P1 should give the same R-values as the better one; if so, P1 can be discarded. Try this with and without twin refinement - although it's hard to compare R-values of non-twinned and twinned refinements. The automatic way to do this is with Zanuda. If you run that locally, you can make refmac do twin refinement. For all resulting structures, I'd also feed the resulting Fcalc (!) into pointless. That should reveal that the packing is indeed close to P3221. Best wishes, Kay On Thu, 26 Aug 2021 11:54:06 +0200, Peer Mittl wrote: >Der CCP4 community, > >Is there a refinement program that can handle protein monomers sitting >on crystallographic 2-folds? > >This is probably a strange question but we have the following situation. >We have a 2.6 Ang datasets in SG P3221 (Rpim=4%, Isa=19.6) and a clear >molrep solution with 2 chains, albeit with tNCS (0/0/0.5) that can be >refined to around 27/33% Rfactor. According to Vm a third chain could be >present. So far so good, but there is clear difference ED for a third >chain sitting exactly on the 2-fold. Since the protein has a peculiar >shape, one can tell even its orientation. I can relax the symmetry to >P32 (or even P1) and place the missing chain with 50% occupancy on the >2-fold. This model can be refined, but I do not like this work around, >because the data is clearly P3221. > >Any hints on similar crystal pathologies and how they have been handled >would be helpful. > >All the best, >Peer > > > >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/ 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/ 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/
[ccp4bb] chain on 2-fold axis?
Der CCP4 community, Is there a refinement program that can handle protein monomers sitting on crystallographic 2-folds? This is probably a strange question but we have the following situation. We have a 2.6 Ang datasets in SG P3221 (Rpim=4%, Isa=19.6) and a clear molrep solution with 2 chains, albeit with tNCS (0/0/0.5) that can be refined to around 27/33% Rfactor. According to Vm a third chain could be present. So far so good, but there is clear difference ED for a third chain sitting exactly on the 2-fold. Since the protein has a peculiar shape, one can tell even its orientation. I can relax the symmetry to P32 (or even P1) and place the missing chain with 50% occupancy on the 2-fold. This model can be refined, but I do not like this work around, because the data is clearly P3221. Any hints on similar crystal pathologies and how they have been handled would be helpful. All the best, Peer 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] unknown density in 1.6 A structure
Dear Nick, I like your idea and your comments on this substance. 1,2-hexandiol supposed to be chiral but probably it is used as a racemic mixture in cosmetics. There is even a crystal structure in the PDB (4EUS) and an associated 3 letter code. I´ll give it a try because it seems to fit nicely the ED map. I just have to find out how to handle racemics in buster refinement. Thank you all for your helpful comments. Have a nice day. -Peer Am 03.06.2021 um 13:00 schrieb Nicholas Keep: How about an overlay of the structure you have with a second ligand conformation where the OH and CH2OH are swapped? ie two alternative conformations of the ligand. That might remove the last green density 1,2-hexanediol is apparently widely used in cosmetics and moisturisers. Did someone somehow touch the protein or crystallisation tray with recently moisturised hands? Even so it must have very high affinity for your protein Best wishes Nick 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/
[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://www.jiscmail.ac.uk/cgi-bin/webadmin?SUBED1=CCP4BB=1
[ccp4bb] Density questionable?
Dear Colleagues, We are working on a structure where the density for a whole protein chain (>200 aa) is questionable, since the B-factors exceed 200 Å2 (2.3 Ang resolution). However, the initial difference density map and the feature enhanced map (normal 2fo-fc map to a minor extend) support the presence of this chain. Putting the chain seems equally wrong as not putting it. Putting it reduces Rfree by 0.3%. As a conservative researcher I feel tempted to deposit the structure without this highly mobile/weakly occupied chain, but other researchers may say "he has missed something". Handling this chain like a weakly occupied water is probably wrong, but what is the optimal/correct way? Is there a general opinion on how the escape this dilemma? All the best, Peer To unsubscribe from the CCP4BB list, click the following link: https://www.jiscmail.ac.uk/cgi-bin/webadmin?SUBED1=CCP4BB=1
Re: [ccp4bb] unidentified crescent-shaped electron density
I have seen similar density with a fatty acid (C16 or C18). In our case the there was also a lysine side chain forming a salt bridge with the carboxylate. Furthermore, the pocket was very hydrophobic. What about this one? -Peer On 02.04.2019 03:12, Zhen Luo wrote: Hi everyone, Thank you very much for your help and time. A PEG fragment indeed seems to fit into the density. Thanks again! Best regards, Zhen *From: *CCP4 bulletin board on behalf of Zhen Luo *Reply-To: *Zhen Luo *Date: *Tuesday, 2 April 2019 at 9:01 AM *To: *"CCP4BB@JISCMAIL.AC.UK" *Subject: *[ccp4bb] unidentified crescent-shaped electron density Dear all, Could you please shed some light on what this crescent-shaped density around the lysine side chain might belong to? I now have two unrelated protein structures where this kind of density can be found surrounding a lysine side chain. Protein 1cid:image009.png@01D4E932.5B911B60 Protein 2cid:image012.png@01D4E932.5B911B60 2FOFC maps were contoured at around 1.5 sigma, FOFC map at 3 sigma. One protein was crystallised in 0.1 M CaCl_2 and 20% PEG 3350; the other in 10% PEG 2, 20% PEG MME 550, 0.03 M CaCl_2 /MgCl_2 , 0.1 M MES/imidazole. Protein buffers contained 0.025 M HEPES and 0.15 M NaCl. None of these fitted in well. Could it be cleaved PEG? Any suggestion would be greatly appreciated. Thanks in advance! Best regards, Zhen Luo School of Chemistry and Molecular Biosciences The University of Queensland, Australia 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 -- **** Peer Mittl, PD Dr. Biochemisches Institut Universität Zürich Room 44M03 Winterthurer Strasse 190 CH-8057 Zürich Tel. +41-(0)44-6356559 Mobile +41-(0)76-2776566 Fax. +41-(0)44-6356805 Mail mi...@bioc.uzh.ch To unsubscribe from the CCP4BB list, click the following link: https://www.jiscmail.ac.uk/cgi-bin/webadmin?SUBED1=CCP4BB=1
[ccp4bb] PhD student position in structure-guided protein engineering at the University of Zürich
We have an immediate opening in the Plückthun lab at the Department of Biochemistry (University of Zürich, Switzerland) for a PhD student to become part of the protein engineering team on the structure-guided development of designed Armadillo-repeat proteins. The lab seeks a talented and motivated individual with expertise in molecular biology, protein purification and characterization with a successfully completed master degree. Previous experience in X-ray crystallography, combinatorial biochemistry and protein engineering would be an asset. Candidates with one or more of these skills with an interest to learn the others are highly encouraged to apply. The project aims to drastically shortcut the protein design cycle by creating constructs of designed Armadillo-repeat proteins (dArmRP) that crystallize predictably, leading to very rapid structure determination. dArmRPs are being developed to develop a general recognition code for unstructured peptide stretches in many scenarios, and the successful candidate would contribute to developing this system. Our lab has an outstanding reputation for the design, characterization, and selection of artificial binding proteins, such as antibodies, and designed ankyrin repeat proteins (DARPins), but so far, all artificial binding proteins must be selected from combinatorial protein libraries. To eliminate the time- and labor-intensive selection process we are establishing dArmRPs as rationally designed binders that specifically recognize the targeted sequence in a predictable way. Background on the dArmRP project can be found in Hansen et al. 2016 (J Am Chem Soc. 138:3526) and 2018 (J Struct Biol. 201:108) and on our homepage (https://www.bioc.uzh.ch/plueckthun/). To support your PhD project we have access to in-house X-ray crystallography facilities, the protein crystallization center and regular access to the Swiss Light Source (SLS), a 3^rd generation synchrotron nearby. We are not only doing cutting-edge basic protein engineering, but we are also applying our tools to target medically relevant questions, particularly from the field of cancer research. We are a very interdisciplinary group with expertise ranging to eukaryotic cell biology and even animal models. This interdisciplinary approach allows a speedy conversion of your results into applied research. The city of Zürich offers a very high standard of living, excellent opportunities for hiking, skiing and aquatic sports, and a thriving arts and culture scene. Please submit your application or informal inquiries to mi...@bioc.uzh.ch or pv...@bioc.uzh.ch. Application deadline: August 15, 2018 To unsubscribe from the CCP4BB list, click the following link: https://www.jiscmail.ac.uk/cgi-bin/webadmin?SUBED1=CCP4BB=1
[ccp4bb] self-rotation in the absence of NCS
We are working with a multi-domain protein crystallized in SG P6_5 with one molecule per asymmetric unit. The structure was refined at 2.00 A resolution with reasonable R-factors but unfortunately the domain we are most interested in seems to be disordered. Interestingly, the self-rotation function shows peaks on the kappa=180° plane (omega=90°, phi=19° (and every 30°)), with more than 50% origin peak height. Therefore, we are wondering if perhaps the space group assignment might be sub-optimal. Any explanations how these self-rotation peaks could occur and how we could extract meaningful information to resolve the disordered domain are welcome. Best regards, Peer P.S. Some additional information: pointless suggests SG P6_5, the data doesn't seem to be twinned (L-test), the refined part of the structure has no internal symmetry and refinement in P1 doesn't reveal the lost domain. -- Peer Mittl, PD Dr. Biochemisches Institut Universität Zürich Room 44M03 Winterthurer Strasse 190 CH-8057 Zürich Tel. +41-(0)44-6356559 Mobile +41-(0)76-2776566 Fax. +41-(0)44-6356834 Mail mi...@bioc.uzh.ch
Re: [ccp4bb] self-rotation in the absence of NCS
Zbyszek Otwinowski and Fred Vellieux suggested to run the self-rotation on Fcalcs. This suggestion solved the problem, since there are similar peaks on the kappa=180° planes as well. However, I wasn't able to get rid of those peaks by playing around with resolution and integration radius. I must say that I am surprized, because - as Eleanor pointed out - I also expected to find peaks on the kappa=180° planes only in case of P6522 symmetry. Anyway, this experience reminds me to run some simple tests beforehand. -Peer On 29.04.2015 15:31, Eleanor Dodson wrote: Well - PG P6/mmm (possible SG P6522) will have peaks at kappa = 180 omega = 90 phi = 0 30 60 etc.. But if there is only one molecule / asymm unit there cant be an extra 2-fold. How big are the relative domains? Your interesting domain couldnt just be cleaved off could it? Eleanor On 29 April 2015 at 12:59, Peer Mittl mi...@bioc.uzh.ch mailto:mi...@bioc.uzh.ch wrote: We are working with a multi-domain protein crystallized in SG P6_5 with one molecule per asymmetric unit. The structure was refined at 2.00 A resolution with reasonable R-factors but unfortunately the domain we are most interested in seems to be disordered. Interestingly, the self-rotation function shows peaks on the kappa=180° plane (omega=90°, phi=19° (and every 30°)), with more than 50% origin peak height. Therefore, we are wondering if perhaps the space group assignment might be sub-optimal. Any explanations how these self-rotation peaks could occur and how we could extract meaningful information to resolve the disordered domain are welcome. Best regards, Peer P.S. Some additional information: pointless suggests SG P6_5, the data doesn't seem to be twinned (L-test), the refined part of the structure has no internal symmetry and refinement in P1 doesn't reveal the lost domain.
Re: [ccp4bb] self-rotation in the absence of NCS
I forgot to acknowledge Pierre Legrand for the same suggestion. -Peer Zbyszek Otwinowski and Fred Vellieux suggested to run the self-rotation on Fcalcs. This suggestion solved the problem, since there are similar peaks on the kappa=180° planes as well. However, I wasn't able to get rid of those peaks by playing around with resolution and integration radius. I must say that I am surprized, because - as Eleanor pointed out - I also expected to find peaks on the kappa=180° planes only in case of P6522 symmetry. Anyway, this experience reminds me to run some simple tests beforehand. -Peer On 29.04.2015 15:31, Eleanor Dodson wrote: Well - PG P6/mmm (possible SG P6522) will have peaks at kappa = 180 omega = 90 phi = 0 30 60 etc.. But if there is only one molecule / asymm unit there cant be an extra 2-fold. How big are the relative domains? Your interesting domain couldnt just be cleaved off could it? Eleanor On 29 April 2015 at 12:59, Peer Mittl mi...@bioc.uzh.ch mailto:mi...@bioc.uzh.ch wrote: We are working with a multi-domain protein crystallized in SG P6_5 with one molecule per asymmetric unit. The structure was refined at 2.00 A resolution with reasonable R-factors but unfortunately the domain we are most interested in seems to be disordered. Interestingly, the self-rotation function shows peaks on the kappa=180° plane (omega=90°, phi=19° (and every 30°)), with more than 50% origin peak height. Therefore, we are wondering if perhaps the space group assignment might be sub-optimal. Any explanations how these self-rotation peaks could occur and how we could extract meaningful information to resolve the disordered domain are welcome. Best regards, Peer P.S. Some additional information: pointless suggests SG P6_5, the data doesn't seem to be twinned (L-test), the refined part of the structure has no internal symmetry and refinement in P1 doesn't reveal the lost domain.
[ccp4bb] Refmac error
Dear Colleagues, For some reason Refmac refuses to read the file mon_lib_list.cif athough the file exists. The error message is as follows: Open failed: Unit: 7, File: /share/prog64/ccp4-6.3.0/lib/data/monomers/list/mon_lib_list.cif (logical: /share/prog64/ccp4-6.3.0/lib/data/monomers/list/mon_lib_list.cif) BFONT COLOR=#FF!--SUMMARY_BEGIN-- Last system error message: Illegal seek Refmac_5.7.0029: Open failed: File: /share/prog64/ccp4-6.3.0/lib/data/monomers/list/mon_lib_list.cif Refmac_5.7.0029: Open failed: File: /share/prog64/ccp4-6.3.0/lib/data/monomers/list/mon_lib_list.cif Any advice would be welcome. -Peer
[ccp4bb] PhD position
At the Department of Biochemistry (University of Zürich) we have an immediate opening for a PhD student working in the field of protein engineering and structural biology. The successful candidate will joint a multidisciplinary team of scientists from combinatorial biochemistry and protein engineering (Prof. A. Plückthun), molecular dynamics (Prof. A. Caflisch), NMR spectroscopy (Prof. O. Zerbe) and X-ray crystallography (PD P. Mittl). It is the goal of this joined project to develop a toolbox of peptide recognition modules (on the basis of the Armadillo repeat) which can be assembled on a rational basis to recognize any extended peptide with a given sequence. Several scaffolds of binding modules have already been developed but it will be the goal of this PhD thesis to design and synthesize artificial Armadillo-repeat proteins that can be crystallized with the cognate polypeptides. Experience (at the master-level) in structure-based planning of mutants, general molecular biology tools, protein expression and purification is mandatory and knowledge of protein X-ray crystallography would be an asset. The project will involve mutant design and structure determination by X-ray crystallography. For further information, please consult the following publications or send an e-mail to either mi...@bioc.uzh.ch or plueckt...@bioc.uzh.ch. Designed Armadillo repeat proteins: library generation, characterization and selection of peptide binders with high specificity. Varadamsetty G, Tremmel D, Hansen S, Parmeggiani F, Plückthun A.J Mol Biol. 2012; 424:68-87 Optimization of designed armadillo repeat proteins by molecular dynamics simulations and NMR spectroscopy. Alfarano P, Varadamsetty G, Ewald C, Parmeggiani F, Pellarin R, Zerbe O, Plückthun A, Caflisch A. Protein Sci. 2012; 21:1298-314 Structure-based optimization of designed Armadillo-repeat proteins. Madhurantakam C, Varadamsetty G, Grütter MG, Plückthun A, Mittl PR. Protein Sci. 2012;21:1015-28 Designed armadillo repeat proteins as general peptide-binding scaffolds: consensus design and computational optimization of the hydrophobic core. Parmeggiani F, Pellarin R, Larsen AP, Varadamsetty G, Stumpp MT, Zerbe O, Caflisch A, Plückthun A.J Mol Biol. 2008 ;376:1282-304
[ccp4bb] P-CUBE workshop on mammalian expression technologies in Oxford
On behalf of the P-cube management team: -Peer Dear All, Don't miss out on the P-CUBE workshop in Oxford in April 2011! Register today under www.p-cube.eu and learn everything about mammalian expression technologies. The registration deadline is March 19, 2011. See you in Oxford! P-CUBE Management Team *** P-CUBE Workshop on Mammalian Expression Technologies Oxford, 3-8 April 2011 The course is designed primarily for structural biologists who have prokaryotic expression experience and would like to use the mammalian expression system for more challenging targets. The goals of the course are to allow participants to have hands-on experience of mammalian cell culture, small and large scale transient gene expression including the use of automated systems, protein purification and crystallization. The course will be structured so as to interleave lectures and practical sessions. The participants will have the opportunity to closely interact with the tutors and to directly discuss their research projects with the experts in small workgroups. PARTICIPANTS Participants will be asked to submit their CV and a letter of motivation. The course will be limited to 12 participants from European structural biology laboratories. • There is no registration fee for the workshop. • Accommodation (5 nights max.) during the workshop will be covered. • Participants are responsible for their own travel costs. TOPICS: 1. Mammalian expression platform overview 2. Mammalian cell lines for protein expression and glycosylation control 3. Mammalian expression constructs design 4. Small scale expression screen with Western Blot 5. Large scale expression and protein purification 6. Protein crystallization 7. Protein structure case studies SPEAKERS: Radu Aricescu (Oxford University, UK), Yuguang Zhao (Oxford University, UK), Joop van den Heuvel (Helmholtz-Zentrum für Infektionsforschung, DE), Veronica Chang (Oxford University, UK), Ben Bishop (Oxford University, UK), Karl Harlos (Oxford University, UK), Tom Walter (Oxford University, UK), Christian Siebold (Oxford University, UK), Joerg Standfuss (Paul Scherrer Institut, CH)
[ccp4bb] P-CUBE - Last chance to sign up for the Workshop and 1st Annual Meeting in Grenoble- Registration is closing soon!
Dear All, Don't miss out on the P-CUBE meeting in Grenoble in September! Sign up today underwww.p-cube.eu. The registration deadline is June 30. 2010. See you in Grenoble P-CUBE Management Team *** Dr. Jutta Tatzel Program Manager P-CUBE Department of Biochemistry University of Zurich Winterthurerstr. 190 CH-8057 Zurich Tel +41 44 635 5593 Email:j.tat...@bioc.uzh.ch
[ccp4bb] P-CUBE: Workshop and 1st Annual Meeting in Grenoble- Registration is closing soon!
On behalf of the P-cube Management Team Dear All, To make sure you don't miss out on the P-CUBE meeting in Grenoble in September, please sign up as soon as possible under*www.p-cube.eu*. The registration deadline is June 30. 2010. See you in Grenoble P-CUBE Management Team
[ccp4bb] Pcube User meeting
Dear Colleagues, I would like to draw your attention to the upcoming Pcube-user meeting (September 8-9, 2010 in Grenoble). You are probably aware that the EU-founded Pcube project provides open access to several supportive technologies for macromolecular crystallography, ranging from fragment screening to advanced microscopy technologies. The best way to learn more about these technologies would be to visit *http://www.p-cube.eu/* and sign up for the meeting. Participation is free of charge but registration is mandatory and the deadline is approaching very quickly. Best regards, Peer
[ccp4bb] Postdoctoral position at UZH on designed repeat proteins
Applications are invited for two vacant positions at the University of Zürich (Switzerland), Department of Biochemistry, group of Prof. Markus Grütter, one at the postdoctoral and one at a research assistant level. We are focussing on the investigation of the structure/function relationship of proteins involved in apoptosis, innate immunity and trans-membrane signalling (PNAS (2008) 105:20251, Structure (2008) 16:1443, Structure (2007) 15:625, PLoS Biol (2007) 5:e7) as well as on protein design of binding proteins such as DARPINs (Structure Review (2008) 16:1443). We seek enthusiastic team players with experience in at least one of the following disciplines: protein X-ray crystallography, protein expression and purification, and eukaryotic expression systems. Good knowledge of English is mandatory. Candidates applying for the Post-Doc position should hold a PhD in a life science-related subject. All positions are time limited for three years. Salary will be according to the regulations of the Swiss National Science Foundation. Please submit your electronic application including a CV, a list of publications (for the Post-Doc positions) and the names of two to three referees to Mrs. Salome Rittmeyer at the following address: secgruet...@bioc.uzh.ch We offer a stimulating and cooperative working environment at an internationally renowned institution. Further information about the Department of Biochemistry and the University of Zürich are available under: http://www.bioc.uzh.ch/index.php?id=3L=1 -- Peer Mittl, PD Dr. Biochemisches Institut Universität Zürich Winterthurer Strasse 190 CH-8057 Zürich Tel. +41-(0)44-635 6559 Fax. +41-(0)44-635 6834 Mail mi...@bioc.uzh.ch
[ccp4bb] 21st Rhine-Knee Regiomeeting 2007
Registration for the 21st Rhine-Knee Regiomeeting 2007 in Hölstein (CH) is open until August 31th, 2007 under http://www.bioc.uzh.ch/index.php?204L=1 . Best regards, Peer Mittl
