Hi Mark, I also use dis.embl and IUPred in Jalview when trying to come up with construct boundaries.
In the specific case I was referring to, we had a secreted human protein with a long and atypical secretion tag, which made it difficult to place the N-terminus of the mature protein within the sequence. My initial attempts didn't work at all, and after reviewing the AF2 model at the EBI database, I removed a further ~30 residues at the N-terminus, which yielded a well expressed, soluble, secreted product. Speaking more generally, I think that AF2 is probably "just another tool" for helping to inform construct boundaries. I have no empirical evidence that the AF2 pLDDT is more sensitive or accurate than other bespoke disorder prediction software. Because it worked once for me, I'll run it/check the EBI database every time from now on, but you're absolutely right that I probably would have done that anyway! Best, Dave -- Dr David C. Briggs CSci MRSB Senior Laboratory Research Scientist Signalling and Structural Biology Lab The Francis Crick Institute London, UK == about.me/david_briggs ________________________________ From: Mark J. van Raaij <mjvanra...@cnb.csic.es> Sent: 05 April 2022 14:47 To: CCP4BB@jiscmail.ac.uk <CCP4BB@JISCMAIL.AC.UK> Cc: David Briggs <david.bri...@crick.ac.uk> 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 predictors? i.e. http://dis.embl.de<https://eur03.safelinks.protection.outlook.com/?url=http%3A%2F%2Fdis.embl.de%2F&data=04%7C01%7C%7C84f2f05bbd5a4598ff5908da170ae108%7C4eed7807ebad415aa7a99170947f4eae%7C0%7C1%7C637847632751920405%7CUnknown%7CTWFpbGZsb3d8eyJWIjoiMC4wLjAwMDAiLCJQIjoiV2luMzIiLCJBTiI6Ik1haWwiLCJXVCI6Mn0%3D%7C2000&sdata=2FjXOEwioG4hdIuef0nPFEM%2B4pV6V%2FB9kYa9phWu4Ds%3D&reserved=0> , which we've used for construct design https://prdos.hgc.jp<https://eur03.safelinks.protection.outlook.com/?url=https%3A%2F%2Fprdos.hgc.jp%2F&data=04%7C01%7C%7C84f2f05bbd5a4598ff5908da170ae108%7C4eed7807ebad415aa7a99170947f4eae%7C0%7C1%7C637847632751920405%7CUnknown%7CTWFpbGZsb3d8eyJWIjoiMC4wLjAwMDAiLCJQIjoiV2luMzIiLCJBTiI6Ik1haWwiLCJXVCI6Mn0%3D%7C2000&sdata=ckpin7cUBV3sVYnc1Coxe72%2FQNMd1M%2ByXpoRaCbdRgE%3D&reserved=0> , which I haven't really tried yet https://iupred2a.elte.hu<https://eur03.safelinks.protection.outlook.com/?url=http%3A%2F%2Fiupred2a.elte.hu%2F&data=04%7C01%7C%7C84f2f05bbd5a4598ff5908da170ae108%7C4eed7807ebad415aa7a99170947f4eae%7C0%7C1%7C637847632751920405%7CUnknown%7CTWFpbGZsb3d8eyJWIjoiMC4wLjAwMDAiLCJQIjoiV2luMzIiLCJBTiI6Ik1haWwiLCJXVCI6Mn0%3D%7C2000&sdata=5IVfLHj7g9Wq4BiQdoez1wB0mgsvQ608xAcw3LzY%2BnM%3D&reserved=0> (which a colleague here likes because it also tries to predict likelihood of protein interactions) Or perhaps it's just because you were going to run alphafold2 anyway? best wishes, Mark Mark J van Raaij Dpto de Estructura de Macromoleculas, lab 20B Centro Nacional de Biotecnologia - CSIC calle Darwin 3 E-28049 Madrid, Spain tel. +34 91 585 4616 (internal 432092) On 4 Apr 2022, at 21:30, David Briggs <david.bri...@crick.ac.uk<mailto:david.bri...@crick.ac.uk>> wrote: 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. Briggs Senior Laboratory Research Scientist Signalling and Structural Biology Lab The Francis Crick Institute London, UK == about.me/david_briggs<https://eur03.safelinks.protection.outlook.com/?url=http%3A%2F%2Fabout.me%2Fdavid_briggs&data=04%7C01%7C%7C84f2f05bbd5a4598ff5908da170ae108%7C4eed7807ebad415aa7a99170947f4eae%7C0%7C1%7C637847632751920405%7CUnknown%7CTWFpbGZsb3d8eyJWIjoiMC4wLjAwMDAiLCJQIjoiV2luMzIiLCJBTiI6Ik1haWwiLCJXVCI6Mn0%3D%7C2000&sdata=RqGd8%2Fy%2FuaLk%2B9IjCfiagj%2BZiHQB%2BYwAu269u7C0HFs%3D&reserved=0> ________________________________ From: CCP4 bulletin board <CCP4BB@JISCMAIL.AC.UK<mailto:CCP4BB@JISCMAIL.AC.UK>> on behalf of Scott Classen <sclas...@lbl.gov<mailto:sclas...@lbl.gov>> Sent: Monday, April 4, 2022 8:06:38 PM To: CCP4BB@JISCMAIL.AC.UK<mailto:CCP4BB@JISCMAIL.AC.UK> <CCP4BB@JISCMAIL.AC.UK<mailto:CCP4BB@JISCMAIL.AC.UK>> Subject: [ccp4bb] Has anyone successfully used RoseTTAFold or AF2 to guide crystallization? External Sender: Use caution. Hello CCP4, Has anyone successfully used the available ML/AI protein folding tools to guide crystallization construct design? Maybe you had a protein or domain that was resistant to crystallization efforts and the folding algorithms predicted some loops or termini that were disordered? Then you trimmed or modified them in some way to aid in crystallization? Or if you haven’t done this yourself, are you aware of anyone who has? Thanks, Scott ~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~ Scott Classen, Ph.D. ALS-ENABLE TomAlberTron Beamline 8.3.1 SIBYLS Beamline 12.3.1 Advanced Light Source Lawrence Berkeley National Laboratory 1 Cyclotron Rd MS6R2100 Berkeley, CA 94720 mobile 510.206.4418 desk 510.495.2697 beamline 510.495.2134 ~~~~~~~~~~~~~~~~~~~~~~~~~~~~~~ ________________________________ To unsubscribe from the CCP4BB list, click the following link: https://www.jiscmail.ac.uk/cgi-bin/WA-JISC.exe?SUBED1=CCP4BB&A=1<https://eur03.safelinks.protection.outlook.com/?url=https%3A%2F%2Fwww.jiscmail.ac.uk%2Fcgi-bin%2FWA-JISC.exe%3FSUBED1%3DCCP4BB%26A%3D1&data=04%7C01%7C%7C84f2f05bbd5a4598ff5908da170ae108%7C4eed7807ebad415aa7a99170947f4eae%7C0%7C1%7C637847632751920405%7CUnknown%7CTWFpbGZsb3d8eyJWIjoiMC4wLjAwMDAiLCJQIjoiV2luMzIiLCJBTiI6Ik1haWwiLCJXVCI6Mn0%3D%7C2000&sdata=Thftvzk1yXUEG0ZM9nIKNUrWKFUu%2FCTj9PcBwy1sMyE%3D&reserved=0> The Francis Crick Institute Limited is a registered charity in England and Wales no. 1140062 and a company registered in England and Wales no. 06885462, with its registered office at 1 Midland Road London NW1 1AT ________________________________ To unsubscribe from the CCP4BB list, click the following link: https://www.jiscmail.ac.uk/cgi-bin/WA-JISC.exe?SUBED1=CCP4BB&A=1<https://eur03.safelinks.protection.outlook.com/?url=https%3A%2F%2Fwww.jiscmail.ac.uk%2Fcgi-bin%2FWA-JISC.exe%3FSUBED1%3DCCP4BB%26A%3D1&data=04%7C01%7C%7C84f2f05bbd5a4598ff5908da170ae108%7C4eed7807ebad415aa7a99170947f4eae%7C0%7C1%7C637847632751920405%7CUnknown%7CTWFpbGZsb3d8eyJWIjoiMC4wLjAwMDAiLCJQIjoiV2luMzIiLCJBTiI6Ik1haWwiLCJXVCI6Mn0%3D%7C2000&sdata=Thftvzk1yXUEG0ZM9nIKNUrWKFUu%2FCTj9PcBwy1sMyE%3D&reserved=0> The Francis Crick Institute Limited is a registered charity in England and Wales no. 1140062 and a company registered in England and Wales no. 06885462, with its registered office at 1 Midland Road London NW1 1AT ######################################################################## To unsubscribe from the CCP4BB list, click the following link: https://www.jiscmail.ac.uk/cgi-bin/WA-JISC.exe?SUBED1=CCP4BB&A=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/
