Dear Xavier, dear all, good points indeed. I guess the aim is to refine an atomic model against a cryo-EM map so that the atomic model describes the experimental data well, aiming for a good final geometry, refined B-factors etc. of the atomic model. Here is a description of what we recently did for one of our structures: https://www.nature.com/protocolexchange/protocols/6365
HTH, Bruno From: CCP4 bulletin board [mailto:[email protected]] On Behalf Of Israel Sanchez Sent: Friday, August 31, 2018 2:32 PM To: [email protected] Subject: Re: [ccp4bb] cryo-EM Dear Xavier, On top of Rob's excellent suggestion, I would recoment you to have a look at this paper by Alan Brown and co-workers (including Rob himself): https://www.ncbi.nlm.nih.gov/pubmed/25615868 We normally performs an initial real-space refinement with phenix.real-space followed by two step of reciprocal space refinement with Refmac. In the first Refmac run we normally use secondary structure restrains generated by ProSmart for proteins and LibG for nuclei acids. In the second Refmac run, we try to release the restrains power and also play around with the matrix governing the weight between the experimental data and ideal symmetry. You can monitor when are you entering over-fitting conditions employing the half maps normally output by Relion (or the program you use for cryoEM data processing). More specific examples here: https://www.ncbi.nlm.nih.gov/pubmed/24675956 and here: https://www.ncbi.nlm.nih.gov/pubmed/24792965 Finally, I would not recommend at all playing around with the phases of a cryoEM map. If you have a region which is less visible due to poorer local resolution, you may blur the map computing a post-precessed map with a lower Bfactor that the one automatically assigned by Relion or low pass filter the same map a bit. Or even combine both approaches. Not sure what the big guys in the field think about tinkering with the phases of a cryoEM map but I feel this is still highly controversial. May be Garib or Pavel think differently? Hope this help. El vie., 31 ago. 2018 a las 8:02, Rob Nicholls (<[email protected]<mailto:[email protected]>>) escribió: Dear Xavier, I recommend having a look at a few of the articles that were published in the special issue of Acta Cryst D from June this year - Proceedings of the third CCP-EM Spring Symposium. https://journals.iucr.org/d/issues/2018/06/00/ Best regards, Rob ------ Original Message ------ From: "F.Xavier Gomis-Rüth" <[email protected]<mailto:[email protected]>> To: [email protected]<mailto:[email protected]> Sent: 31/08/2018 12:52:48 Subject: [ccp4bb] cryo-EM Dear all, is there some kind of general trend if not consensus as how to refine cryo-EM structures? What do people do for maps in the 3-4Å range, pure RSR against the untouched map and that's it? Reciprocal space refinement against the backtransformed structure factors from the map but without touching the map? I've even heard that some people "dare" to calculate 2mFo-DFc-maps.... Thanks a lot for your feedback. Best, Xavier -- [cid:[email protected]] ________________________________ To unsubscribe from the CCP4BB list, click the following link: https://www.jiscmail.ac.uk/cgi-bin/webadmin?SUBED1=CCP4BB&A=1 ________________________________ To unsubscribe from the CCP4BB list, click the following link: https://www.jiscmail.ac.uk/cgi-bin/webadmin?SUBED1=CCP4BB&A=1 -- Israel S. Fernandez PhD Ramakrishnan-Lab MRC Laboratory of Molecular Biology, Francis Crick Avenue, Cambridge Biomedical Campus, Cambridge CB2 0QH, UK. ________________________________ To unsubscribe from the CCP4BB list, click the following link: https://www.jiscmail.ac.uk/cgi-bin/webadmin?SUBED1=CCP4BB&A=1 ######################################################################## To unsubscribe from the CCP4BB list, click the following link: https://www.jiscmail.ac.uk/cgi-bin/webadmin?SUBED1=CCP4BB&A=1
