Re: [ccp4bb] Please share your experience about "ugly" crystals showing good diffraction
I couldn’t agree more with Patrick which is often the case. To echo some of his comments, in the High-Throughput Crystallization Screening Center we see many examples where the visual images of initial crystallization hits are very poor and experienced (or inexperienced) crystallographers would typically ignore them. SONICC and UV two photon fluorescence images can indicate something far more promising. Optimization of the visually ‘crappy’ crystals (or even things that really don’t look like crystals to the eye) that have strong SONICC or UV two photon fluorescence very often produces beautiful results. We published a short paper on this “The detection and subsequent volume optimization of biological nanocrystals, Luft JR, et al.. Struct Dyn. 2015 May 15;2(4):041710 and have seen many examples since. The crystallization research page of the High-Throughput Screening Center (http://getacrystal.org) has a link to this paper under the Crystallization Research section and there is a more extensive literature related to this research are on the website in my signature. Excuse the shameless plug, but if you don’t have SONICC and UV-two photon detection, this is a standard part of the crystallization screening center available to all and run by Dr. Sarah Bowman (https://hwi.buffalo.edu/scientist-directory/sbowman/) – Details at http://getacrystal.org. There is also some news on automated outcome classification just to put a teaser of things to come out there. Best, Eddie Edward Snell Ph.D. Biological Small Angle Scattering Theory and Practice, Eaton E. Lattman, Thomas D. Grant, and Edward H. Snell. Available through all good bookshops, or direct from Oxford University Press Director of the NSF BioXFEL Science and Technology Center President and CEO Hauptman-Woodward Medical Research Institute BioInnovations Chaired Professorship, University at Buffalo, SUNY 700 Ellicott Street, Buffalo, NY 14203-1102 hwi.buffalo.edu Phone: (716) 898 8631 Fax: (716) 898 8660 Skype:eddie.snell Email: esn...@hwi.buffalo.edu Webpage: https://hwi.buffalo.edu/scientist-directory/snell/ [cid:image001.png@01D4150E.CF9BD9D0] Heisenberg was probably here! From: CCP4 bulletin board [mailto:CCP4BB@JISCMAIL.AC.UK] On Behalf Of Patrick Shaw Stewart Sent: Friday, July 6, 2018 7:08 AM To: CCP4BB@JISCMAIL.AC.UK Subject: Re: [ccp4bb] Please share your experience about "ugly" crystals showing good diffraction Hi All I have a comment, based on my old supervisor's explanation, which seemed to make sense. Crystals usually grow layer by layer. Once a new layer is formed it quickly expands to cover the whole surface. That's why crystals normally have flat surfaces and sharp edges - the layers/steps expand rapidly until they get to the edges. However it doesn't have to be like that. Sometimes new layers can form roughly as quickly as the previous layers can spread. The result is crystals with curved surfaces - or even just blobs. Just because the new layers form at a rate that is comparable to the spreading doesn't mean that the crystals won't be ordered, and won't diffract well. Once I understood that I understood what I was seeing better when I checked my drops. Best wishes Patrick On 5 July 2018 at 22:06, Sanishvili, Ruslan mailto:rsanishv...@anl.gov>> wrote: Hi Anirban, It would be great if you could share the compilation of relevant responses to your request. I think many others in the community could use these examples for educational purposes. Best, Nukri Ruslan Sanishvili (Nukri), Ph.D. Macromolecular Crystallographer GM/CA@APS X-ray Science Division, ANL 9700 S. Cass Ave. Lemont, IL 60439 Tel: (630)252-0665 Fax: (630)252-0667 rsanishv...@anl.gov<mailto:rsanishv...@anl.gov> From: CCP4 bulletin board mailto:CCP4BB@JISCMAIL.AC.UK>> on behalf of Anirban Banerjee mailto:ani...@gmail.com>> Sent: Thursday, June 28, 2018 7:07 PM To: CCP4BB@JISCMAIL.AC.UK<mailto:CCP4BB@JISCMAIL.AC.UK> Subject: [ccp4bb] Please share your experience about "ugly" crystals showing good diffraction Dear all, Apologies for the non-CCP4 related question. If you have concrete experience about visually unappealing, i.e. ugly crystals ( your own take on ugly is fine) diffracting better than comparably similar sized nicer looking crystals of the same protein, will you please share here ? Even better if that led to a published structure. Might you also have pictures ? We have all heard anecdotes about not using visual appearance to judge the quality of crystals as far as their ability to give good diffraction data is concerned but I am trying to gather some concrete pointers here to motivate trainees. Thanks very much for any help. Best, Anirban P.S. I know that there is probably a lot of thought and wisdom on this this specific t
Re: [ccp4bb] Please share your experience about "ugly" crystals showing good diffraction
Dear Anirban, On 05/07/18 22:06, Sanishvili, Ruslan wrote: Hi Anirban, It would be great if you could share the compilation of relevant responses to your request. I think many others in the community could use these examples for educational purposes. Quite so, and I am sure that a lot of people on this list will agree with me that there was no need to apologise for asking this question ;-) Regards, Peter. *From:* CCP4 bulletin board on behalf of Anirban Banerjee *Sent:* Thursday, June 28, 2018 7:07 PM *To:* CCP4BB@JISCMAIL.AC.UK *Subject:* [ccp4bb] Please share your experience about "ugly" crystals showing good diffraction Dear all, Apologies for the non-CCP4 related question. If you have concrete experience about visually unappealing, i.e. ugly crystals ( your own take on ugly is fine) diffracting better than comparably similar sized nicer looking crystals of the same protein, will you please share here ? Even better if that led to a published structure. Might you also have pictures ? We have all heard anecdotes about not using visual appearance to judge the quality of crystals as far as their ability to give good diffraction data is concerned but I am trying to gather some concrete pointers here to motivate trainees. Thanks very much for any help. Best, Anirban P.S. I know that there is probably a lot of thought and wisdom on this this specific topic but I am really looking for actual experience. 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 -- Peter Keller Tel.: +44 (0)1223 353033 Global Phasing Ltd., Fax.: +44 (0)1223 366889 Sheraton House, Castle Park, Cambridge CB3 0AX United Kingdom To unsubscribe from the CCP4BB list, click the following link: https://www.jiscmail.ac.uk/cgi-bin/webadmin?SUBED1=CCP4BB=1
Re: [ccp4bb] Please share your experience about "ugly" crystals showing good diffraction
Hi All I have a comment, based on my old supervisor's explanation, which seemed to make sense. Crystals usually grow layer by layer. Once a new layer is formed it quickly expands to cover the whole surface. That's why crystals normally have flat surfaces and sharp edges - the layers/steps expand rapidly until they get to the edges. However it doesn't have to be like that. Sometimes new layers can form roughly as quickly as the previous layers can spread. The result is crystals with curved surfaces - or even just blobs. Just because the new layers form at a rate that is comparable to the spreading doesn't mean that the crystals won't be ordered, and won't diffract well. Once I understood that I understood what I was seeing better when I checked my drops. Best wishes Patrick On 5 July 2018 at 22:06, Sanishvili, Ruslan wrote: > Hi Anirban, > > It would be great if you could share the compilation of relevant responses > to your request. I think many others in the community could use these > examples for educational purposes. > > Best, > > Nukri > > > Ruslan Sanishvili (Nukri), Ph.D. > Macromolecular Crystallographer > GM/CA@APS > X-ray Science Division, ANL > 9700 S. Cass Ave. > Lemont, IL 60439 > > Tel: (630)252-0665 > Fax: (630)252-0667 > rsanishv...@anl.gov > > > > -- > *From:* CCP4 bulletin board on behalf of Anirban > Banerjee > *Sent:* Thursday, June 28, 2018 7:07 PM > *To:* CCP4BB@JISCMAIL.AC.UK > *Subject:* [ccp4bb] Please share your experience about "ugly" crystals > showing good diffraction > > > Dear all, > > Apologies for the non-CCP4 related question. > > If you have concrete experience about visually unappealing, i.e. ugly > crystals ( your own take on ugly is fine) diffracting better than > comparably similar sized nicer looking crystals of the same protein, will > you please share here ? Even better if that led to a published structure. > Might you also have pictures ? > > We have all heard anecdotes about not using visual appearance to judge the > quality of crystals as far as their ability to give good diffraction data > is concerned but I am trying to gather some concrete pointers here to > motivate trainees. > > Thanks very much for any help. > > Best, > > Anirban > > P.S. I know that there is probably a lot of thought and wisdom on this > this specific topic but I am really looking for actual experience. > > -- > > 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 > -- patr...@douglas.co.ukDouglas Instruments Ltd. Douglas House, East Garston, Hungerford, Berkshire, RG17 7HD, UK Directors: Peter Baldock, Patrick Shaw Stewart http://www.douglas.co.uk Tel: 44 (0) 148-864-9090US toll-free 1-877-225-2034 Regd. England 2177994, VAT Reg. GB 480 7371 36 To unsubscribe from the CCP4BB list, click the following link: https://www.jiscmail.ac.uk/cgi-bin/webadmin?SUBED1=CCP4BB=1
Re: [ccp4bb] Please share your experience about "ugly" crystals showing good diffraction
Hi Anirban, It would be great if you could share the compilation of relevant responses to your request. I think many others in the community could use these examples for educational purposes. Best, Nukri Ruslan Sanishvili (Nukri), Ph.D. Macromolecular Crystallographer GM/CA@APS X-ray Science Division, ANL 9700 S. Cass Ave. Lemont, IL 60439 Tel: (630)252-0665 Fax: (630)252-0667 rsanishv...@anl.gov From: CCP4 bulletin board on behalf of Anirban Banerjee Sent: Thursday, June 28, 2018 7:07 PM To: CCP4BB@JISCMAIL.AC.UK Subject: [ccp4bb] Please share your experience about "ugly" crystals showing good diffraction Dear all, Apologies for the non-CCP4 related question. If you have concrete experience about visually unappealing, i.e. ugly crystals ( your own take on ugly is fine) diffracting better than comparably similar sized nicer looking crystals of the same protein, will you please share here ? Even better if that led to a published structure. Might you also have pictures ? We have all heard anecdotes about not using visual appearance to judge the quality of crystals as far as their ability to give good diffraction data is concerned but I am trying to gather some concrete pointers here to motivate trainees. Thanks very much for any help. Best, Anirban P.S. I know that there is probably a lot of thought and wisdom on this this specific topic but I am really looking for actual experience. To unsubscribe from the CCP4BB list, click the following link: https://www.jiscmail.ac.uk/cgi-bin/webadmin?SUBED1=CCP4BB=1 To unsubscribe from the CCP4BB list, click the following link: https://www.jiscmail.ac.uk/cgi-bin/webadmin?SUBED1=CCP4BB=1
Re: [ccp4bb] Please share your experience about "ugly" crystals showing good diffraction
Good evening, Over the years I've seen great diffraction from any number of hideous brittle plates with perforated edges, 'Krypton skyscrapers', curved whiskers, 'hypodermic needles', even from crystals that looked like cat vomit (hairballs), and so forth. The structure *ab ovo* story is my favorite, though: Similar to Debanu's case - we've had crystals (of a novel insecticidal toxin) that were perfectly egg-shaped, and every one of them was the same shape - regardless of differences in their size. I mounted them on a bet with my colleagues and we were cracking all kinds of jokes about making omelets with X-rays, and so forth - right until the very first egg diffracted better than 1.6A and produced a lovely structure. On top of this bit of weirdness, the only way these crystals could be grown was by reductive methylation followed by proteolysis. Opposite order of steps did not work, and each step in itself did not work. This story brings to mind the hellish experience of polishing small molecule crystals down to little balls in order to minimize absorption effects (great, now I made myself feel old). Artem - Cosmic Cats approve of this message On Thu, Jun 28, 2018 at 8:07 PM, Anirban Banerjee wrote: > > Dear all, > > Apologies for the non-CCP4 related question. > > If you have concrete experience about visually unappealing, i.e. ugly > crystals ( your own take on ugly is fine) diffracting better than > comparably similar sized nicer looking crystals of the same protein, will > you please share here ? Even better if that led to a published structure. > Might you also have pictures ? > > We have all heard anecdotes about not using visual appearance to judge the > quality of crystals as far as their ability to give good diffraction data > is concerned but I am trying to gather some concrete pointers here to > motivate trainees. > > Thanks very much for any help. > > Best, > > Anirban > > P.S. I know that there is probably a lot of thought and wisdom on this > this specific topic but I am really looking for actual experience. > > -- > > To unsubscribe from the CCP4BB list, click the following link: > https://www.jiscmail.ac.uk/cgi-bin/webadmin?SUBED1=CCP4BB=1 > To unsubscribe from the CCP4BB list, click the following link: https://www.jiscmail.ac.uk/cgi-bin/webadmin?SUBED1=CCP4BB=1
Re: [ccp4bb] Please share your experience about "ugly" crystals showing good diffraction
Hi Anirban, Never thought someone will ask for image of 'ugly' crystals, but fortunately I have found some saved pictures which is more than 8 years old! I think I have a perfect example. The pictures can be found in the folder by following the link: https://drive.google.com/drive/folders/0B1M5Wq4437-CN0pLWXdOUG44MlE?usp=sharing These ugly crystals diffracted to 1.95 Å as opposed to some nice looking crystals (grown in different condition) that diffracted to quite lower resolution. The structure from these crystals have PDB ID is 2XQ0 ( https://www.rcsb.org/structure/2XQ0). The structure is described in JMB journal ( https://www.sciencedirect.com/science/article/pii/S0022283610012969?via%3Dihub ). Best wishes, Mahmudul Lund Sweden On Fri, Jun 29, 2018 at 8:57 AM Janet Newman wrote: > Hi Anirban, > > Single easy example to find (see attached images) - gorgeous crystals - > sub 3A (grown in sodium malonate) > Lozenge crystals - better than 2A (grown in ammonium sulfate) > > (Barbiturase protein ) > > Cheers, Janet > > -Original Message- > From: CCP4 bulletin board [mailto:CCP4BB@JISCMAIL.AC.UK] On Behalf Of > Debanu Das > Sent: Friday, 29 June 2018 4:36 PM > To: CCP4BB@JISCMAIL.AC.UK > Subject: Re: [ccp4bb] Please share your experience about "ugly" crystals > showing good diffraction > > Hi Anirban, > > At JCSG, we subjected ~180,000 crystals from ~3500 unique/novel > proteins/complexes to X-ray diffraction screening, resulting in >1500 > novel structures in the PDB at an average resolution ~2.0A. In theory, > if we had the bandwidth now to sort through all that data to pull out > images of the mounted crystals and their diffraction quality, we could > probably get some good analysis done on this. > > However, as an alternative, I can certainly provide some examples that > we have dealt with recently that I hope will add substance to the > anecdotes to motivate your trainees! > > a) For one protein, our first crystals were oval shaped, without sharp > geometry/edges, much like a pebble. We got a high quality 1.9A > structure out of it. Subsequent optimization led to much nicer looking > crystals with nice shapes and we collected ~50 data sets from them all > resulting in ~2A structures so not much improved compared to the > initial visually poorer crystals. > > b) For another protein, crystals grow nicely but some of the crystals > remain suspended in solution whereas other settle to the bottom and > stick. For the ones which stick, some can be dislodged by gentle > prodding with a nylon loop while harvesting and they result in (along > with the ones that are not stuck) in ~1.8-2A structures. For the ones > which are stuck and cannot be gently dislodged, a nudge with a plastic > tip (out of desperation!) is sufficient to dislodge them, and they > retain their nice visual appearance and shape, but have total loss in > diffraction. > > c) We see this too for crystals soaked with ligands. In some cases > after soaking the crystals appear fine but suffer in diffraction > quality, and in some cases they appear to have suffered visually but > result in usable data sets/structures. > > So at the end of the day there are crystals that appear nice but do > not diffract well and there are crystals that do not appear nice but > lead to good/usable structures. So it's all about inner beauty. Never > give up on crystals/crystal optimization without testing them by > X-rays. > > Best, > Debanu > > On Thu, Jun 28, 2018 at 5:07 PM, Anirban Banerjee > wrote: > > > > Dear all, > > > > Apologies for the non-CCP4 related question. > > > > If you have concrete experience about visually unappealing, i.e. ugly > > crystals ( your own take on ugly is fine) diffracting better than > > comparably similar sized nicer looking crystals of the same protein, will > > you please share here ? Even better if that led to a published structure. > > Might you also have pictures ? > > > > We have all heard anecdotes about not using visual appearance to judge > the > > quality of crystals as far as their ability to give good diffraction > data is > > concerned but I am trying to gather some concrete pointers here to > motivate > > trainees. > > > > Thanks very much for any help. > > > > Best, > > > > Anirban > > > > P.S. I know that there is probably a lot of thought and wisdom on this > this > > specific topic but I am really looking for actual experience. > > > > > > > > To unsubscribe from the CCP4BB list, click the following link: > > https://www.jiscmail.ac.uk/cgi-bin/webadmin?SUBED1=CCP4BB=1 > > #
Re: [ccp4bb] Please share your experience about "ugly" crystals showing good diffraction
Hi Anirban, At JCSG, we subjected ~180,000 crystals from ~3500 unique/novel proteins/complexes to X-ray diffraction screening, resulting in >1500 novel structures in the PDB at an average resolution ~2.0A. In theory, if we had the bandwidth now to sort through all that data to pull out images of the mounted crystals and their diffraction quality, we could probably get some good analysis done on this. However, as an alternative, I can certainly provide some examples that we have dealt with recently that I hope will add substance to the anecdotes to motivate your trainees! a) For one protein, our first crystals were oval shaped, without sharp geometry/edges, much like a pebble. We got a high quality 1.9A structure out of it. Subsequent optimization led to much nicer looking crystals with nice shapes and we collected ~50 data sets from them all resulting in ~2A structures so not much improved compared to the initial visually poorer crystals. b) For another protein, crystals grow nicely but some of the crystals remain suspended in solution whereas other settle to the bottom and stick. For the ones which stick, some can be dislodged by gentle prodding with a nylon loop while harvesting and they result in (along with the ones that are not stuck) in ~1.8-2A structures. For the ones which are stuck and cannot be gently dislodged, a nudge with a plastic tip (out of desperation!) is sufficient to dislodge them, and they retain their nice visual appearance and shape, but have total loss in diffraction. c) We see this too for crystals soaked with ligands. In some cases after soaking the crystals appear fine but suffer in diffraction quality, and in some cases they appear to have suffered visually but result in usable data sets/structures. So at the end of the day there are crystals that appear nice but do not diffract well and there are crystals that do not appear nice but lead to good/usable structures. So it's all about inner beauty. Never give up on crystals/crystal optimization without testing them by X-rays. Best, Debanu On Thu, Jun 28, 2018 at 5:07 PM, Anirban Banerjee wrote: > > Dear all, > > Apologies for the non-CCP4 related question. > > If you have concrete experience about visually unappealing, i.e. ugly > crystals ( your own take on ugly is fine) diffracting better than > comparably similar sized nicer looking crystals of the same protein, will > you please share here ? Even better if that led to a published structure. > Might you also have pictures ? > > We have all heard anecdotes about not using visual appearance to judge the > quality of crystals as far as their ability to give good diffraction data is > concerned but I am trying to gather some concrete pointers here to motivate > trainees. > > Thanks very much for any help. > > Best, > > Anirban > > P.S. I know that there is probably a lot of thought and wisdom on this this > specific topic but I am really looking for actual experience. > > > > 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
[ccp4bb] Please share your experience about "ugly" crystals showing good diffraction
Dear all, Apologies for the non-CCP4 related question. If you have concrete experience about visually unappealing, i.e. ugly crystals ( your own take on ugly is fine) diffracting better than comparably similar sized nicer looking crystals of the same protein, will you please share here ? Even better if that led to a published structure. Might you also have pictures ? We have all heard anecdotes about not using visual appearance to judge the quality of crystals as far as their ability to give good diffraction data is concerned but I am trying to gather some concrete pointers here to motivate trainees. Thanks very much for any help. Best, Anirban P.S. I know that there is probably a lot of thought and wisdom on this this specific topic but I am really looking for actual experience. To unsubscribe from the CCP4BB list, click the following link: https://www.jiscmail.ac.uk/cgi-bin/webadmin?SUBED1=CCP4BB=1
