I have not much to contribute to this discussion, but I did run three refinements - TLS as defined in the PDb, Iso B factors only, and unrestrained aniso Bs . The free R factors are almost the same for all three, Rwork same for TLS and IsoBs - lower for aniso.. I checked the peak heights for the anomalous scattering Zn in all three anom diff maps, and the allBiso was the highest ( 15 sigma v 12 Sigma for the others..) Is that significant? probably not! All models and maps overlapped very closely of course. Eleanor Dodson
On Wed, 30 Oct 2024 at 10:45, Italo Carugo Oliviero < olivieroitalo.car...@unipv.it> wrote: > Dear Robbie, > > thank you for this very detailed and inspiring contribution to the > discussion. > > However, three key issues must be taken into account when dividing a > protein structure into numerous small miniblocks. It is crucial to remember > that these miniblocks must remain rigid, with their interatomic distances > kept constant. This is because not only must the R-factor significantly > decrease (Hamilton), but the final stereochemistry (including x, y, z > coordinates, and B-factors) must also be coherent. > > (i) Ideally, the definition of miniblocks should be grounded in a sound > theoretical framework based on chemical and physical principles. > > (ii) If the definition of miniblocks relies on internal data rather than a > theoretical basis, it is essential to ensure reproducibility, meaning the > same miniblocks should be consistently defined using different crystals, > different diffraction data collected by different researchers, etc. > > (iii) Moreover, it would be beneficial to assess the rigidity of these > miniblocks through methods such as MD or NMR, whether in the solid state or > in solution. > > Otherwise, I will remain a ‘Sceptical Chymist’, to quote Robert Boyle's > famous book (1661, subtitle: ‘Chymico-Physical Doubts & Paradoxes’). And > the reliability of B-factors produced with these miniblocks TLS > refinements, even if published in important biology journals, runs the risk > of being comparable to adventures of Harry Potter, to quote a series of > books much more widely read than Boyle's book. > > Best, > > Oliviero > > Il giorno mar 29 ott 2024 alle ore 17:34 Robbie Joosten < > robbie_joos...@hotmail.com> ha scritto: > >> Hear, hear! Although I like the Latin expression and the occasional rant >> in general, I fully agree with Gerlind's point. >> >> TLS refinement is a great tool and it can help even if the model >> segmentation is not biologically relevant. However, not biologically >> relevant may not necessarily mean that the model is overfitted. This is >> what we test in PDB-REDO: we take all defined TLS models (typically the >> model in the coordinate file and one-group-per-chain, but you can specify >> mode models) and refine with these. Then we use a version of the Hamilton >> test to see if the improved fit with the data comes from a genuine >> improvement of the model rather than from just adding extra parameters. >> This approach has flagged quite a few cases in the PDB where the TLS >> segmentation is a bit "overenthusiastic". However in the case of 8q66 the >> more complex model is, from a refinement point-of-view, fine. I haven't >> read the paper for this structure model, but if the authors are not >> deriving any specific conclusions from the TLS model, I do not see it as a >> problem. >> >> As Gerlind points out, we should teach the right things. In this case >> there are a few things users should know: >> - TLS refinement is often a means to an end, not a (biological) model >> analysis. >> - TLS analysis (i.e. grouping refined (anisotropic) B-factors to fit a >> domain-like model of "motion") on the other hand is a useful tool if done >> carefully. >> - Do not read too much into a given TLS model if you do not know where it >> came from. Also don't if you do. >> - Don't overparameterise your TLS model, apply Occam's razor, and test >> for overfitting. >> - You can check your TLS model with tools like Parvati. Perhaps having >> more validation tools would be nice. >> >> Cheers, >> Robbie >> >> > -----Original Message----- >> > From: CCP4 bulletin board <CCP4BB@JISCMAIL.AC.UK> On Behalf Of Gerlind >> > Sulzenbacher >> > Sent: Tuesday, October 29, 2024 17:08 >> > To: CCP4BB@JISCMAIL.AC.UK >> > Subject: Re: [ccp4bb] Structures refined ad mentula canis (this is >> Latin) >> > >> > Dear Italo, >> > >> > It seems to me that the ccp4bb has always been a very gentle and >> empathic >> > platform of exchange of ideas and offering a great deal of help to non- >> > experts, "ignorants" as your friend calls them. >> > >> > I remember that in the late 90thies there were some ayatollahs, which >> have >> > become gentle civils by now, that used accusatory citations to underpin >> their >> > claims. This period is over, or at least I hope so. >> > >> > >> > If ignorance exists, it's our fault of not being able to teach properly >> the young >> > generation. >> > >> > I understand your frustation and may be it deserves a letter in an Acta >> > Journal, but please, please, refrain of citing the "ignorants" in this >> bb. >> > >> > Wishing you a happy retirement and nice and fruitfuil projects, other >> than B- >> > factors, for your future. >> > >> > Gerlind >> > >> > >> > >> > >> > >> > On 29/10/2024 16:38, Italo Carugo Oliviero wrote: >> > >> > >> > Ce mail provient de l'extérieur, restons vigilants >> > >> > In a few months, I will retire, because my brain thinks it is >> still young >> > but the rest of the body disagrees and thinks the brain is slightly >> senile. Of >> > course, I will miss the protein structures very much, so unpredictable >> in their >> > insolent beauty. >> > >> > >> > >> > >> > I have devoted several years of my life to the B-factors of their >> atoms. >> > >> > >> > >> > >> > Initially, almost 30 years ago, some colleagues mocked me, >> telling me >> > that the B-side of proteins was nothing but dust hidden under the >> carpet, >> > nothing but background noise. Some later changed their minds, and it is >> now >> > generally agreed that the B-side of proteins deserves to be visited and >> > admired. >> > >> > >> > >> > >> > However, I am concerned about more recent developments that are >> > turning B-factors into real junk. TLS refinements as audacious as they >> are >> > ridiculously unprofessional are producing disasters. For example, the >> 8q66 >> > structure is very strange. It has been split into individual TLS >> mini-blocks, >> > which clearly demonstrates that the authors are unaware of the TLS >> > procedure and its proper application. A protein containing two >> structural >> > domains was divided into six small TLS blocks, one of which belongs to >> both >> > domains. Where are physics and chemistry? >> > >> > >> > >> > How can this happen? Here is the answer from a friend and >> > colleague: “unfortunately, the crystallographic programs became so >> > automatic and easy to use nowadays, that even complete ignorant people >> > can get something from them, and do not think if what they got makes >> sense >> > or not.” >> > >> > >> > >> > >> > >> > >> > Are computational models preferable in the end? >> > >> > >> > ________________________________ >> > >> > To unsubscribe from the CCP4BB list, click the following link: >> > https://www.jiscmail.ac.uk/cgi-bin/WA- >> > JISC.exe?SUBED1=CCP4BB&A=1 >> > >> > >> > >> > >> > -- >> > Gerlind Sulzenbacher >> > Architecture et Fonction des Macromolécules Biologiques >> > UMR7257 CNRS, Aix-Marseille Université >> > Case 932 >> > 163 Avenue de Luminy >> > 13288 Marseille cedex 9 >> > France >> > Tel +33 413 94 95 27 >> > E-mail: gerlind.sulzenbac...@univ-amu.fr >> > <mailto:gerlind.sulzenbac...@univ-amu.fr> >> > >> > ________________________________ >> > >> > >> > To unsubscribe from the CCP4BB list, click the following link: >> > https://www.jiscmail.ac.uk/cgi-bin/WA-JISC.exe?SUBED1=CCP4BB&A=1 >> >> >> ######################################################################## >> >> 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/ >> > > ------------------------------ > > To unsubscribe from the CCP4BB list, click the following link: > https://www.jiscmail.ac.uk/cgi-bin/WA-JISC.exe?SUBED1=CCP4BB&A=1 > ######################################################################## 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/
