Re: [ccp4bb] [3dem] [ccpem] Which resolution?
Hi Marin, crystallography has long moved away from the term 'resolution', see e.g. https://www.cell.com/structure/fulltext/S0969-2126(18)30138-2. It is merely a ballpark number, and it is good to know whether crystallographic data were cut at 1, 2, or 3 Angstrom, but not very important. What counts is the interpretation of the model and conclusion that can be drawn on the system under study. It requires a broader understanding of crystallography in order to understand whether the conclusions are justified. Resolution plays only a minor role in this. It is more useful to take a look at the crystallographic map itself in order to understand. EM is totally different from crystallography, and why would one mix concepts between the fields? Best, Tim On Thursday, February 13, 2020 12:07:15 AM CET Marin van Heel wrote: > Hi Tim, > Good to hear from you! No longer at PSI?? > See... You are already touching upon one of the logical breaking points in > the resolutiton story...! X-ray crystallography resolution criteria like > R-factors make absolutely no sense outside the field of crystallography and > of structural biology. It is the result of a hybrid iterative optimisation > process between the phases of a model structure and the measured amplitudes > of a diffraction experiment! The FRC/FSC resolution criteria, in contrast, > are universal quality metrics not at all coupled to Cryo-EM or structural > biology. Using structural biology arguments like how well I see an alpha > helix or how well I see the hole in an aromatic ring as an assessment > criterion of whether a metric is good or not is a waste of time! (Moreover > filtering a map can completley change its appearance without changing its > information contents). Even some my own (ex-)students and (ex-)postdocs > sometimes completely miss this fundamental point. The FRC and FSC criteria > are now used as quality metrics in all walks of image science like X-ray > tomography and super-resolution light microscopy, fields of science where > atomic coordinates of proteins are not an issue. The FRC / FSC functions > are universal and very direct metrics that compare both the amplitudes and > the phases of two independent measurements of images or 3D-densities of the > same object. For more details, see the 2017 bioRxiv paper and references > therein (https://www.biorxiv.org/content/10.1101/224402v1) and check my > #WhyOWhy tweets (@marin_van_heel). See also: van Heel - Unveiling ribosomal > structures: the final phases - Current opinion in structural biology 10 > (2000) 259-264. > > Cheers, > Marin > > On Wed, Feb 12, 2020 at 11:22 AM Tim Gruene wrote: > > Dear Marin, > > > > I did not read the enire thread, nor the manuscript you point at - > > apologize > > in case this has been discussed before. > > > > What about a practical approach to determine the resolution of a cryoEM > > map: > > one could take a feature with scales of interest, e.g. an alpha-helix, and > > shift and/or rotate it in steps of, say, 0.3A in several directions to > > see, at > > which magnitude (degree / distance) refinement does not take the helix > > back to > > its original position (within error margins). > > > > One could also take a Monte-Carlo approach and do an arbitrary number of > > random re-orientations of such a helix, refine, and calculate the > > variation in > > position and rotation. > > > > This would reflect my understanding of resolution, much more than any > > statistical descriptor. > > > > Best regards, > > Tim > > > > On Wednesday, February 12, 2020 1:46:48 PM CET Marin van Heel wrote: > > > Hi Laurence, > > > > > > One thing is certain: the 0.143 threshold is RUBBISH and all CC50 etc > > > are > > > also based on the same SLOPPY STATISTICS as are all fixed-valued FSC > > > thresholds. This controversy has been ragings for a long long time and > > > > the > > > > > errors made were extensively described (again) in our most recent paper > > > (Van Heel & Schatz 2017 BioRxiv: > > > https://www.biorxiv.org/content/10.1101/224402v1) which has been > > > > downloaded > > > > > more than 3000 times. Further papers on the issue are in the pipeline. > > > > The > > > > > math BLUNDER behind this controversy is simple: the inner product > > > > between > > > > > a signal vector and a noise vector is NOT zero (but rather proportional > > > > to > > > > > SQRT(N) where N is the length of the vectors) and cannot be left out of > > > > the > > > > > equations. This error goes back to a paper published in Nature in 1975 > > > > and > > > > > has since been repeated frequently, including in the first paper > > > > promoting > > > > > the erroneous 0.143 FSC threshold. The consequences of this blunder in > > > current processing are serious especially when these erroneous metrics > > > > are > > > > > used as an optimisation criterion in iterative refinements at > > > resolutions > > > close to Nyquist. I get tired of facing this s
Re: [ccp4bb] [3dem] [ccpem] Which resolution?
Dear Pavel, Your paper is one of the more elaborate ones on the issue with an exhaustive list of references! No wonder, since some 20 years ago, Bruno Klaholtz was a very successful post-doc in my group at Imperial in London. However, I have discussed this paper with Bruno at our Brazil School in 2018 and I pointed out to him that your use of fixed-valued FSC thresholds makes that your paper, like 95% of the papers on resolution in our field, implicitly is based on the "sloppy statistics" of others, namely that the inner product between signal and noise vectors cannot be neglected. As long as those "sloppy statistics" papers are not taken from the literature, years or decades after they have been refuted, and the criticism against them is simply ignored or belittled, incorrect follow-up research along the same sloppy pathways is the consequence, and that is so sad... Sorry, Marin On Wed, Feb 12, 2020 at 8:34 PM Pavel Afonine wrote: > Interesting conversation! I see the 2017 paper is on bioRxiv. I wonder if > it ever made into a peer reviewed journal (couldn't find quickly)? > @Tim Gruene : have a look at d_model in > https://www.ncbi.nlm.nih.gov/pubmed/30198894 which is sort of along > similar lines of what you are hinting here. > Pavel > > On Wed, Feb 12, 2020 at 3:07 PM Marin van Heel < > 057a89ab08a1-dmarc-requ...@jiscmail.ac.uk> wrote: > >> Hi Tim, >> Good to hear from you! No longer at PSI?? >> See... You are already touching upon one of the logical breaking points >> in the resolutiton story...! X-ray crystallography resolution criteria >> like R-factors make absolutely no sense outside the field of >> crystallography and of structural biology. It is the result of a hybrid >> iterative optimisation process between the phases of a model structure and >> the measured amplitudes of a diffraction experiment! The FRC/FSC >> resolution criteria, in contrast, are universal quality metrics not at all >> coupled to Cryo-EM or structural biology. Using structural biology >> arguments like how well I see an alpha helix or how well I see the hole in >> an aromatic ring as an assessment criterion of whether a metric is good or >> not is a waste of time! (Moreover filtering a map can completley change >> its appearance without changing its information contents). Even some my own >> (ex-)students and (ex-)postdocs sometimes completely miss this fundamental >> point. The FRC and FSC criteria are now used as quality metrics in all >> walks of image science like X-ray tomography and super-resolution light >> microscopy, fields of science where atomic coordinates of proteins are not >> an issue. The FRC / FSC functions are universal and very direct metrics >> that compare both the amplitudes and the phases of two independent >> measurements of images or 3D-densities of the same object. For more >> details, see the 2017 bioRxiv paper and references therein ( >> https://www.biorxiv.org/content/10.1101/224402v1) and check my #WhyOWhy >> tweets (@marin_van_heel). See also: van Heel - Unveiling ribosomal >> structures: the final phases - Current opinion in structural biology 10 >> (2000) 259-264. >> >> Cheers, >> Marin >> >> >> On Wed, Feb 12, 2020 at 11:22 AM Tim Gruene >> wrote: >> >>> Dear Marin, >>> >>> I did not read the enire thread, nor the manuscript you point at - >>> apologize >>> in case this has been discussed before. >>> >>> What about a practical approach to determine the resolution of a cryoEM >>> map: >>> one could take a feature with scales of interest, e.g. an alpha-helix, >>> and >>> shift and/or rotate it in steps of, say, 0.3A in several directions to >>> see, at >>> which magnitude (degree / distance) refinement does not take the helix >>> back to >>> its original position (within error margins). >>> >>> One could also take a Monte-Carlo approach and do an arbitrary number of >>> random re-orientations of such a helix, refine, and calculate the >>> variation in >>> position and rotation. >>> >>> This would reflect my understanding of resolution, much more than any >>> statistical descriptor. >>> >>> Best regards, >>> Tim >>> >>> On Wednesday, February 12, 2020 1:46:48 PM CET Marin van Heel wrote: >>> > Hi Laurence, >>> > >>> > One thing is certain: the 0.143 threshold is RUBBISH and all CC50 etc >>> are >>> > also based on the same SLOPPY STATISTICS as are all fixed-valued FSC >>> > thresholds. This controversy has been ragings for a long long time and >>> the >>> > errors made were extensively described (again) in our most recent paper >>> > (Van Heel & Schatz 2017 BioRxiv: >>> > https://www.biorxiv.org/content/10.1101/224402v1) which has been >>> downloaded >>> > more than 3000 times. Further papers on the issue are in the pipeline. >>> The >>> > math BLUNDER behind this controversy is simple: the inner product >>> between >>> > a signal vector and a noise vector is NOT zero (but rather >>> proportional to >>> > SQRT(N) where N is the length of the vectors) and cannot be left out >
Re: [ccp4bb] [3dem] [ccpem] Which resolution?
Interesting conversation! I see the 2017 paper is on bioRxiv. I wonder if it ever made into a peer reviewed journal (couldn't find quickly)? @Tim Gruene : have a look at d_model in https://www.ncbi.nlm.nih.gov/pubmed/30198894 which is sort of along similar lines of what you are hinting here. Pavel On Wed, Feb 12, 2020 at 3:07 PM Marin van Heel < 057a89ab08a1-dmarc-requ...@jiscmail.ac.uk> wrote: > Hi Tim, > Good to hear from you! No longer at PSI?? > See... You are already touching upon one of the logical breaking points in > the resolutiton story...! X-ray crystallography resolution criteria like > R-factors make absolutely no sense outside the field of crystallography and > of structural biology. It is the result of a hybrid iterative optimisation > process between the phases of a model structure and the measured amplitudes > of a diffraction experiment! The FRC/FSC resolution criteria, in contrast, > are universal quality metrics not at all coupled to Cryo-EM or structural > biology. Using structural biology arguments like how well I see an alpha > helix or how well I see the hole in an aromatic ring as an assessment > criterion of whether a metric is good or not is a waste of time! (Moreover > filtering a map can completley change its appearance without changing its > information contents). Even some my own (ex-)students and (ex-)postdocs > sometimes completely miss this fundamental point. The FRC and FSC criteria > are now used as quality metrics in all walks of image science like X-ray > tomography and super-resolution light microscopy, fields of science where > atomic coordinates of proteins are not an issue. The FRC / FSC functions > are universal and very direct metrics that compare both the amplitudes and > the phases of two independent measurements of images or 3D-densities of the > same object. For more details, see the 2017 bioRxiv paper and references > therein (https://www.biorxiv.org/content/10.1101/224402v1) and check my > #WhyOWhy tweets (@marin_van_heel). See also: van Heel - Unveiling > ribosomal structures: the final phases - Current opinion in structural > biology 10 (2000) 259-264. > > Cheers, > Marin > > > On Wed, Feb 12, 2020 at 11:22 AM Tim Gruene > wrote: > >> Dear Marin, >> >> I did not read the enire thread, nor the manuscript you point at - >> apologize >> in case this has been discussed before. >> >> What about a practical approach to determine the resolution of a cryoEM >> map: >> one could take a feature with scales of interest, e.g. an alpha-helix, >> and >> shift and/or rotate it in steps of, say, 0.3A in several directions to >> see, at >> which magnitude (degree / distance) refinement does not take the helix >> back to >> its original position (within error margins). >> >> One could also take a Monte-Carlo approach and do an arbitrary number of >> random re-orientations of such a helix, refine, and calculate the >> variation in >> position and rotation. >> >> This would reflect my understanding of resolution, much more than any >> statistical descriptor. >> >> Best regards, >> Tim >> >> On Wednesday, February 12, 2020 1:46:48 PM CET Marin van Heel wrote: >> > Hi Laurence, >> > >> > One thing is certain: the 0.143 threshold is RUBBISH and all CC50 etc >> are >> > also based on the same SLOPPY STATISTICS as are all fixed-valued FSC >> > thresholds. This controversy has been ragings for a long long time and >> the >> > errors made were extensively described (again) in our most recent paper >> > (Van Heel & Schatz 2017 BioRxiv: >> > https://www.biorxiv.org/content/10.1101/224402v1) which has been >> downloaded >> > more than 3000 times. Further papers on the issue are in the pipeline. >> The >> > math BLUNDER behind this controversy is simple: the inner product >> between >> > a signal vector and a noise vector is NOT zero (but rather proportional >> to >> > SQRT(N) where N is the length of the vectors) and cannot be left out of >> the >> > equations. This error goes back to a paper published in Nature in 1975 >> and >> > has since been repeated frequently, including in the first paper >> promoting >> > the erroneous 0.143 FSC threshold. The consequences of this blunder in >> > current processing are serious especially when these erroneous metrics >> are >> > used as an optimisation criterion in iterative refinements at >> resolutions >> > close to Nyquist. I get tired of facing this systematic misuse of the >> FSC >> > function, which I myself have introduced into the literature in >> 1982/1986, >> > and people nevertheless feel they know better (with no scientific >> arguments >> > to support!) and they feel justified to use it beyond its definition >> range, >> > and to continue to ignore the correct math. To counter this systematic >> > abuse of my brain child - over decades - I feel the need to use CLEAR >> > LANGUAGE! >> > Have fun! >> > Marin >> >> -- >> -- >> Tim Gruene >> Head of the Centre for X-ray Structure Analysis >> Faculty of Chemistry >> U
Re: [ccp4bb] [3dem] [ccpem] Which resolution?
Hi Tim, Good to hear from you! No longer at PSI?? See... You are already touching upon one of the logical breaking points in the resolutiton story...! X-ray crystallography resolution criteria like R-factors make absolutely no sense outside the field of crystallography and of structural biology. It is the result of a hybrid iterative optimisation process between the phases of a model structure and the measured amplitudes of a diffraction experiment! The FRC/FSC resolution criteria, in contrast, are universal quality metrics not at all coupled to Cryo-EM or structural biology. Using structural biology arguments like how well I see an alpha helix or how well I see the hole in an aromatic ring as an assessment criterion of whether a metric is good or not is a waste of time! (Moreover filtering a map can completley change its appearance without changing its information contents). Even some my own (ex-)students and (ex-)postdocs sometimes completely miss this fundamental point. The FRC and FSC criteria are now used as quality metrics in all walks of image science like X-ray tomography and super-resolution light microscopy, fields of science where atomic coordinates of proteins are not an issue. The FRC / FSC functions are universal and very direct metrics that compare both the amplitudes and the phases of two independent measurements of images or 3D-densities of the same object. For more details, see the 2017 bioRxiv paper and references therein (https://www.biorxiv.org/content/10.1101/224402v1) and check my #WhyOWhy tweets (@marin_van_heel). See also: van Heel - Unveiling ribosomal structures: the final phases - Current opinion in structural biology 10 (2000) 259-264. Cheers, Marin On Wed, Feb 12, 2020 at 11:22 AM Tim Gruene wrote: > Dear Marin, > > I did not read the enire thread, nor the manuscript you point at - > apologize > in case this has been discussed before. > > What about a practical approach to determine the resolution of a cryoEM > map: > one could take a feature with scales of interest, e.g. an alpha-helix, and > shift and/or rotate it in steps of, say, 0.3A in several directions to > see, at > which magnitude (degree / distance) refinement does not take the helix > back to > its original position (within error margins). > > One could also take a Monte-Carlo approach and do an arbitrary number of > random re-orientations of such a helix, refine, and calculate the > variation in > position and rotation. > > This would reflect my understanding of resolution, much more than any > statistical descriptor. > > Best regards, > Tim > > On Wednesday, February 12, 2020 1:46:48 PM CET Marin van Heel wrote: > > Hi Laurence, > > > > One thing is certain: the 0.143 threshold is RUBBISH and all CC50 etc are > > also based on the same SLOPPY STATISTICS as are all fixed-valued FSC > > thresholds. This controversy has been ragings for a long long time and > the > > errors made were extensively described (again) in our most recent paper > > (Van Heel & Schatz 2017 BioRxiv: > > https://www.biorxiv.org/content/10.1101/224402v1) which has been > downloaded > > more than 3000 times. Further papers on the issue are in the pipeline. > The > > math BLUNDER behind this controversy is simple: the inner product > between > > a signal vector and a noise vector is NOT zero (but rather proportional > to > > SQRT(N) where N is the length of the vectors) and cannot be left out of > the > > equations. This error goes back to a paper published in Nature in 1975 > and > > has since been repeated frequently, including in the first paper > promoting > > the erroneous 0.143 FSC threshold. The consequences of this blunder in > > current processing are serious especially when these erroneous metrics > are > > used as an optimisation criterion in iterative refinements at resolutions > > close to Nyquist. I get tired of facing this systematic misuse of the > FSC > > function, which I myself have introduced into the literature in > 1982/1986, > > and people nevertheless feel they know better (with no scientific > arguments > > to support!) and they feel justified to use it beyond its definition > range, > > and to continue to ignore the correct math. To counter this systematic > > abuse of my brain child - over decades - I feel the need to use CLEAR > > LANGUAGE! > > Have fun! > > Marin > > -- > -- > Tim Gruene > Head of the Centre for X-ray Structure Analysis > Faculty of Chemistry > University of Vienna > > Phone: +43-1-4277-70202 > > GPG Key ID = A46BEE1A > To unsubscribe from the CCP4BB list, click the following link: https://www.jiscmail.ac.uk/cgi-bin/webadmin?SUBED1=CCP4BB&A=1
