Hi Everyone, Look up J Mol Biol 270 724-738 (Valegard et al.), or doi https://doi.org/10.1006/jmbi.1997.1144 , PDB entry 1ZDH, where the M&M section describes crystallising MS2 virus at 37C. From memory, as they were collecting data at Daresbury during my time there, the crystals were in a gel, after bringing the set up to RT. It helped them a great deal, because then they could cut out the gel and orient the crystals favourably from knowledge of the morphology. That was way before cryo-cooling was available, and a complete data set collection used to require lots of crystals. It helped to know where the gaps in the rotation range were.
Although I haven’t done any experiment at a temp higher than RT, I expect that if you get crystals at a higher temp, then you would likely have them better ordered as you ‘cool’ them down to RT. There is an added advantage in creature comfort without compromising the precious little crystals. Win-win? Pierre ******************************************************* Dr Pierre Rizkallah, Senior Lecturer Structural Biology Institute of Infection & Immunology, Sir Geraint Evans Building, School of Medicine, Heath Campus, Cardiff, CF14 4XN email: rizkall...@cardiff.ac.uk<mailto:rizkall...@cardiff.ac.uk> phone: +44 29 2074 2248 http://www.cardiff.ac.uk/people/view/126690-rizkallah-pierre From: CCP4 bulletin board <CCP4BB@JISCMAIL.AC.UK> On Behalf Of Newman, Janet (Manufacturing, Parkville) Sent: 01 August 2019 23:30 To: CCP4BB@JISCMAIL.AC.UK Subject: Re: [ccp4bb] Importance of temperature during initial crystallization screening Interesting topic, Certainly the two papers suggested by Georg are relevant, and I fully agree with the comments from Daniel that it is hard to predict the behaviour of any given protein from a statistical analysis of proteins in general. I find it interesting that even with the use of incubators to store and image crystal experiments (which takes away the issue of setting up plates in the cold) we still find that our facility users have a strong bias towards 20C over 8C – for example, our standard initial screen (Shotgun) has been set up just over 360 times in the last year, 216 times at 20C and 135 times at 8C. It is equally easy to type “20” or “8” into the request for the plate storage temperature, so its not ease of crystallisation setup which dictates this. It might well be that the thought of harvesting crystals from cold plates puts people off the lower temperature? A quick search through our database shows that of the shotgun screens that were set up in the last year, 51% of those set up at 20C were (human) scored as containing crystals, and 47% of those set up at 8C were (human) scored as containing crystals. So the rate of crystal formation at the two temperature is essentially the same. Patrick, I can’t comment on “unusual” temperatures, as I don’t have any significant experience with going outside the 4-20 region. Cheers, Janet From: CCP4 bulletin board <CCP4BB@JISCMAIL.AC.UK<mailto:CCP4BB@JISCMAIL.AC.UK>> On Behalf Of Georg Mlynek Sent: Friday, 2 August 2019 5:09 AM To: CCP4BB@JISCMAIL.AC.UK<mailto:CCP4BB@JISCMAIL.AC.UK> Subject: Re: [ccp4bb] Importance of temperature during initial crystallization screening Hi Sergei, this publication should be useful for you. https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4756611/<https://eur03.safelinks.protection.outlook.com/?url=https%3A%2F%2Fwww.ncbi.nlm.nih.gov%2Fpmc%2Farticles%2FPMC4756611%2F&data=01%7C01%7Crizkallahp%40CARDIFF.AC.UK%7Cef05a217ab0648bf03dc08d716cfcf58%7Cbdb74b3095684856bdbf06759778fcbc%7C1&sdata=VMEkokroAJl82%2FxlwA0EkW7qfiMQ0YMCRVjE8vumSyk%3D&reserved=0> Additionally it is proposed that when your protein is not so stable (lower Tm), one should incubate the screens at 4C. http://scripts.iucr.org/cgi-bin/paper?S0907444911036225<https://eur03.safelinks.protection.outlook.com/?url=http%3A%2F%2Fscripts.iucr.org%2Fcgi-bin%2Fpaper%3FS0907444911036225&data=01%7C01%7Crizkallahp%40CARDIFF.AC.UK%7Cef05a217ab0648bf03dc08d716cfcf58%7Cbdb74b3095684856bdbf06759778fcbc%7C1&sdata=cCtTPTN9rt0VS4DKTyP82rEGNcWSfyiQNzTjo%2BBVacY%3D&reserved=0> Br, Georg. Am 2019-08-01 um 2:54 PM schrieb Patrick Shaw Stewart: Hi Sergei We did some data-mining on this way, way back, in 2004. See the second section in this link https://www.douglas.co.uk/PDB_data.htm<https://eur03.safelinks.protection.outlook.com/?url=https%3A%2F%2Fwww.douglas.co.uk%2FPDB_data.htm&data=01%7C01%7Crizkallahp%40CARDIFF.AC.UK%7Cef05a217ab0648bf03dc08d716cfcf58%7Cbdb74b3095684856bdbf06759778fcbc%7C1&sdata=9x5egAU%2FgeBintnHRZfKUu4MSXsDPcW%2BBlgYw3qqeaU%3D&reserved=0> When you consider the non-standard temps - ie NOT 4C or 20C - it looks as though the higher-end temps may work better. But of course it's hard to make sense of the results of a martingale. Thx Patrick PS Janet (Newman) do you have anything more up-to-date on this? On Thu, Aug 1, 2019 at 10:24 AM Sergei Strelkov <sergei.strel...@kuleuven.be<mailto:sergei.strel...@kuleuven.be>> wrote: Dear all, I wondered if someone could point me to a recent study on the importance of temperature during initial search for crystallization conditions. It would be interesting to see any real statistics on this subject. We typically try to perform screening at at two temperatures, such as duplicating a given kit screen at 20C and 4C if there is enough sample. My 'gut feeling' is that this is not as important as sampling the chemical space though. Thank you! Sergei Prof. Sergei V. 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