Hi
Forgive me if I’ve missed something, but I can’t find a way to run AlphaFold
without installing it locally - in which case I need a reasonable GPU.
On the other hand, RoseTTAFold does pretty much the same thing and is available
via David Baker’s web server - upload your sequence and sit back and wait. The
overall models that come out are very similar to those from AlphaFold (not
_quite_ as good…) but are generally available wihtout having to install the
software and maybe invest in a good GPU (DeepMind doesn’t have to worry about
the cost or hassle of such things).
https://www.bakerlab.org/index.php/2021/07/15/accurate-protein-structure-prediction-accessible/
(see the link at the bottom of the page just next to Minkyung Baek’s photo)
On the third hand, both AlphaFold and RoseTTAFold are very good at finding
models for proteins that have good order at the secondary structure level. I
don’t know much about IDPs, but I thought that they were intrinsically
disordered because they lack 3D structure. Secondary structure is most
certainly three dimensional…
Just another two ha’porth.
Harry
> On 17 Aug 2021, at 16:59, 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
>
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