To paraphrase from our 2018 Acta Cryst D74(6) paper: “There has been debate as to whether refinement should be performed in real space or reciprocal space; both approaches have advantages and disadvantages. In real space, refinement can be performed locally, which has advantages for computational speed and parallelisation. [Update: Refmac now has the option to perform “local refinement”, which means focusing on just a localised region in order to speed things up – this option is available from within the CCP-EM interface.] Refinement in reciprocal space allows accounting for errors in Fourier coefficients more accurately. And, although errors in neighbouring Fourier coefficients are correlated, they are less correlated than between proximal points in real space. Furthermore, the degree to which errors are correlated will depend on the nature of the underlying data (i.e. on the reconstruction methods). There is a common misconception that the original data are in real space. Reconstructions are typically performed in Fourier space using the projection-slice theorem, before the reconstructed maps are subsequently calculated in real space. Alternatively, 3D reconstruction may be performed in real space using back-projection, noting that this results in a large correlation radius of errors in real space. The procedures in real and reciprocal space are mathematically equivalent; in both cases refinement should be equivalent (apart from details of implementation). The community would benefit from clarification about best practice on this controversial topic, thus proper analysis will be required in future."
Indeed, until works have been published on the objective comparative analysis of the performance of real vs reciprocal space refinement of models against cryo-EM maps, it seems premature to sit too strongly on one side of the fence or the other. Until then, I'd recommend people to try different tools and use what works best for them, without too much preconception. Regards, Rob Dr Rob Nicholls Senior Investigator Scientist MRC Laboratory of Molecular Biology Francis Crick Avenue Cambridge Biomedical Campus Cambridge CB2 0QH > On 30 Jul 2020, at 13:28, Schreuder, Herman /DE <herman.schreu...@sanofi.com> > wrote: > > Hi everybody, > > Thank you very much for the many tips! I discovered that in Coot by clicking > on the N- and C-terminus of a protein chain, one can indeed run a real-space > refinement on the complete chain and the same is true for a rigid body fit. > However, subsequent refinement with Refmac did not converge, so it looks like > I still have to go manually through the chains. ☹ > Still on my to do list are: > - test Isolde > - test Main > - test the wiggle-fit option in Coot. > > Although the Refmac procedure consisting of converting the cryo-EM map into > structure factors and running a "crystallographic" refinement on it seems to > work, in my opinion the future will be a robust real-space refinement on the > cryo-EM map itself, instead of a refinement on derived structure factors. > However, this is my personal opinion. > > Best regards, > Herman > > -----Ursprüngliche Nachricht----- > Von: CCP4 bulletin board <CCP4BB@JISCMAIL.AC.UK> Im Auftrag von Tom Burnley - > UKRI STFC > Gesendet: Mittwoch, 29. Juli 2020 18:51 > An: CCP4BB@JISCMAIL.AC.UK > Betreff: Re: [ccp4bb] AW: [EXTERNAL] Re: [ccp4bb] real real-space-refinement > > EXTERNAL : Real sender is owner-ccp...@jiscmail.ac.uk > > > > Hi Herman, > > > There is lots of information on CCP-EM's modelling tools here: > > > https://urldefense.proofpoint.com/v2/url?u=https-3A__www.ccpem.ac.uk_training_icknield-5F2019_icknield-5F2019.php&d=DwIFAg&c=Dbf9zoswcQ-CRvvI7VX5j3HvibIuT3ZiarcKl5qtMPo&r=HK-CY_tL8CLLA93vdywyu3qI70R4H8oHzZyRHMQu1AQ&m=aaxzRy8JT0AM0_MqNvU2fr6IB4wIbkY8SsgvQkYcw_w&s=D5VOMK2Fz8AOV1PCvJpD9tm914KUxJlsRNRWXYPb7tI&e= > > > This includes a tutorial on Refmac. As you note this is done in reciprocal > space but the CCP-EM pipeline handles conversions from MRC to MTZ format (and > runs Refmac via CCP4). > > > All the best, > > > Tom > > ________________________________ > From: CCP4 bulletin board <CCP4BB@JISCMAIL.AC.UK> on behalf of Boaz Shaanan > <bshaa...@bgu.ac.il> > Sent: 29 July 2020 16:31:46 > To: ccp4bb > Subject: Re: [ccp4bb] AW: [EXTERNAL] Re: [ccp4bb] real real-space-refinement > > Hi, > Try searching ccp-em. > Boaz > > Boaz Shaanan, Ph.D. > Department of Life Sciences > Ben Gurion University of the Negev > Beer Sheva > Israel > > On Jul 29, 2020 18:29, "Schreuder, Herman /DE" <herman.schreu...@sanofi.com> > wrote: > Coot does it, but if one wants to do it for a complete protein, it is a lot > of clicking. On the other hand, I have not tried to just select the N- and > C-terminus for real-space refinement to see what happens. I would prefer to > have some script to do it. > Best, Herman > > Von: Eleanor Dodson <eleanor.dod...@york.ac.uk> > Gesendet: Mittwoch, 29. Juli 2020 17:23 > An: Schreuder, Herman /DE <herman.schreu...@sanofi.com> > Cc: CCP4BB@JISCMAIL.AC.UK > Betreff: [EXTERNAL] Re: [ccp4bb] real real-space-refinement > > > EXTERNAL : Real sender is > eleanor.dod...@york.ac.uk<mailto:eleanor.dod...@york.ac.uk> > > Wont coot do that? > Eleanor > > On Wed, 29 Jul 2020 at 16:20, Schreuder, Herman /DE > <herman.schreu...@sanofi.com<mailto:herman.schreu...@sanofi.com>> wrote: > Dear BB, > > I would like to do a real real-space-refinement of a protein against a > cryo-EM map; not the mtz-based Refmac approach. A quick internet search > produced a lot of Phenix hits, but little ccp4 hits. 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