What the DeepMind people say about this is that when part of a predicted structure has a low predicted lDDT (a CASP measure of model quality), this is a good indicator that it may be intrinsically disordered.
Best wishes, Randy Read > On 17 Aug 2021, at 17:13, Sorin Draga <sor.dr...@gmail.com> wrote: > > Dear all, > > Regarding the AlphaFold2: while its results are impressive for properly > folded proteins, I would caution against using it for IDPs: they do not have > a stable 3D structure that can be predicted, instead they occupy an ensemble > of structures, with a high degree of heterogeneity. While you will find > models of IDPs predicted by AlphaFold, they only represent (at best) one of > many local minima that these proteins occupy. > > Again, many thanks for all your excellent contributions! > > Kind regards, > > Sorin > > On Tue, Aug 17, 2021 at 11:59 AM George Sheldrick <gshe...@uni-goettingen.de> > wrote: > > > > > > As Joel has suggested before, alphafold on an IDP would be interesting and > would seem like a zero-cost starting point - perhaps one you have tried > already. > > > > Sent from ProtonMail mobile > > > > -------- Original Message -------- > On 15 Aug 2021, 15:53, Scott Horowitz < scott.horow...@du.edu> wrote: > > Hi Sorin, > > I hate to say it, but this is a really tough and expensive one. Solving a > true conformational ensemble of one IDP of decent size (~>70 residues) at > something like decent resolution is hard, and not that many labs actually do > it (it's usually a different set of NMR techniques than solving folding > proteins, and that knowledge is even somewhat specialized even within the NMR > community). Solving a co-structural ensemble of two IDPs that bind is even > harder, and I'm hard pressed to remember a single case right now where it's > been done (probably has, but very rarely). Assuming they express really well > and produce decent spectra, it is in theory doable, but I'd assume multiple > years of work by a very good student or postdoc from a lab that specializes > in this and many thousands of dollars (I'd very roughly assume ~$10k in > materials costs alone) would be required for that co-structure. > > The SAXS route is certainly less expensive and faster if it works and gets > you the info you need, but it certainly will be low-res. I'm not as familiar > with it, but if you can differentially label the proteins, the neutron > equivalent of SAXS might also help with the co-structural ensemble to > differentiate which protein is where in the resulting blob. > > Scott > > Scott Horowitz, Ph.D. > Assistant Professor > Department of Chemistry & Biochemistry > Knoebel Institute for Healthy Aging > University of Denver > > ECS Building > 2155 E. Wesley Ave > Denver, CO 80208 > Phone: 303-871-4326 > Fax: 303-871-7915 > Zoom Room: https://udenver.zoom.us/my/scotthorowitz > Email: scott.horow...@du.edu > Office: Room 561 Lab: Room 505 > > From: CCP4 bulletin board <CCP4BB@JISCMAIL.AC.UK> on behalf of Roopa Thapar > <0000070a21fba45f-dmarc-requ...@jiscmail.ac.uk> > Sent: Sunday, August 15, 2021 8:20 AM > To: CCP4BB@JISCMAIL.AC.UK <CCP4BB@JISCMAIL.AC.UK> > Subject: [EXTERNAL] Re: [ccp4bb] biomolecular NMR for IDPs > > [External Email From]: owner-ccp...@jiscmail.ac.uk > > > Hello Sorin, > > > 1. The cost of getting NMR data on the IDPs you propose depends upon the > expression levels of the protein/s as you will need to label with 15N and 13C > - and depending upon your overall yields per liter of E.coli culture, this > can add up. In addition you will need to run triple resonance experiments - > so you should look into the hourly charge to access the NMR spectrometers > where you are located. Moreover, you need to account for time required for > optimization of solution conditions to collect the NMR data as the sample > needs to be homogenous (as in no aggregation) at millimolar or hundreds of > micromolar concentration. As Ethan Merritt suggested, it would be a good > idea to use SAXS first as it requires very little sample, no isotope > labeling, and you can try to narrow down the solution conditions that would > be best suited for NMR. The Kratky plots, Rg values under different solution > conditions can give very useful information about conformational states and > ensembles populated by IDPs. However, although NMR tends to be more > expensive than other techniques but is perfect for IDPs as you point out you > can get residue specific information. A combined NMR/SAXS approach has > proven to be very useful to validate computational models. > > 2. In general, CROs are much more expensive particularly for generating > isotopically labeled samples - it is cost-prohibitive for academic labs. > Genscript is one CRO that will express proteins, but I am not sure if they > will make isotopically labeled proteins for NMR. > > 3. The amount of protein needed depends upon the size of the molecule. You > will need at least 2-3 samples that are differentially labeled with 15N, 13C > (also since you want data on the free and bound forms of the complex) at 0.5 > - 1 mM depending upon the size of the molecule which relates to the > complexity of the NMR spectrum due to number of resonances and the relaxation > times. The total volume required for each sample is between 280 ul - 600 ul, > depending upon which type of instrumentation and NMR probes you have access > to. > > Hope this helps! > > Best regards, > Roopa > > On Saturday, August 14, 2021, 04:12:58 PM CDT, Sorin Draga > <sor.dr...@gmail.com> wrote: > > > Hello everyone, > > I do realize that this is not a NMR focused group, but I do hope that there > are a few spectroscopists lurking around that could possibly answer a few > questions (I am more of a modeler/computationalist): > > The problem: I have two intrinsically disordered proteins that are known to > interact (let's call them 1 and 2). I would like to get structural > information (a conformational ensemble) for 1 and for the "complex" (1+2). > Further down the line (depending on whether this is possible) I would also > like to evaluate potential small molecule inhibitors for the said complex. > Both 1 and 2 are <200 aminoacids long. > > The questions: > > 1. Could the cost of determining the "structure" for 1 and 1+2 be estimated? > To be more precise, I am looking for a ball-park figure on how much a NMR > measurement would cost in this case. > 2. Could anyone recommend a good group/CRO that could provide such a service > and not have an astronomical cost? > 3. Any other suggestions/thoughts that you think might be worth mentioning > (minimum quantity of protein necessary, purity, type of NMR etc) > > Many thanks for your help and time! > > Cheers! > > Sorin > > 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 > > > 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 > > > 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 > ----- Randy J. Read Department of Haematology, University of Cambridge Cambridge Institute for Medical Research Tel: +44 1223 336500 The Keith Peters Building Fax: +44 1223 336827 Hills Road E-mail: rj...@cam.ac.uk Cambridge CB2 0XY, U.K. www-structmed.cimr.cam.ac.uk ######################################################################## 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/
