[Ifeffit] Beryllium XAFS in proteins

Tue Dec 18 17:17:33 CST 2012

Hi Grant

thanks for your comment.

Aluminium has been considered, rather frustratingly we have been unable to replace the Be with other ions. They do not lead to activation of the protein possibly because of the very specific organization of the Be binding site which likely does not suit other ions accept for at low affinity perhaps.  Aluminium is much larger than Be even though it has similar chemistry we have no  good results testing Al to date because it binds to many of the matrices we use to determine if its bound to the protein, for instance Biacore although  aluminium tests have not led to activation in cell based assays either. This is why Be causes the  disease chronic berylliosis  when bound to this protein, the site is so suited to bind only Be and once in there the Be is impossible to remove, with citrate, EDTA or EGTA, and various other chemical means. There are no recorded data to show that aluminum can cause the same disease profile.

I would have loved the opportunity to use the anomalous signal of aluminum if I could get it in the protein but it still would not fully determine the species of Be bound since the size of aluminium may preclude the proper binding conformation for  Be and hence its species.

Such a frustrating problem….

Gina

On Dec 18, 2012, at 3:59 PM, grant bunker wrote:

> Gina - measuring Be in proteins is extremely difficult and probably unprecedented, but it may be possible to substitute another element that is chemically similar and probe that instead. For example, often metalloenzymes remain functional under such substitutions, e.g. if you substitute a Mn or a Co in a Ca site.
>
> The covalent radius of Be is very small, so substituting Ca might not be feasible without excessively distorting the site, but Aluminum isn't far off, and might work. Of course Aluminum itself isn't easy because of its low k-edge, but it may be feasible.
>
> If a heavier element (Ca or heavier) could be made to substitute and retain the functionality/properties you are studying, it might be a suitable probe.
>
> good luck!
>
> Grant Bunker
>
>
> On Dec 18, 2012, at 3:19 PM, ifeffit-request at millenia.cars.aps.anl.gov wrote:
>
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>> Today's Topics:
>>
>>  1. Re: Beryllium and EXAFS (Erik Farquhar)
>>  2. Larch 0.9.17 available for download and testing (Matt Newville)
>>
>>
>> ----------------------------------------------------------------------
>>
>> Message: 1
>> Date: Tue, 18 Dec 2012 12:50:30 -0800 (PST)
>> From: Erik Farquhar <efarq at yahoo.com>
>> To: XAFS Analysis using Ifeffit <ifeffit at millenia.cars.aps.anl.gov>
>> Subject: Re: [Ifeffit] Beryllium and EXAFS
>> Message-ID:
>>      <1355863830.4845.YahooMailNeo at web141002.mail.bf1.yahoo.com>
>> Content-Type: text/plain; charset="iso-8859-1"
>>
>> A bit unrelated to the question at hand, but regarding #5, yes, people do neutron PX.? The focus?is almost entirely on locating hydrogen atoms (e.g. for protonation state), so I wonder what?the neutron PX guys?would say about looking for a Be atom. The downside is that you usually need?quite large crystals compared to X-ray PX and long data collection times.
>> ?
>> Erik
>>
>>
>> ________________________________
>> From: Matthew Marcus <mamarcus at lbl.gov>
>> To: Erik Farquhar <efarq at yahoo.com>; XAFS Analysis using Ifeffit <ifeffit at millenia.cars.aps.anl.gov>
>> Sent: Tuesday, December 18, 2012 1:39 PM
>> Subject: Re: [Ifeffit] Beryllium and EXAFS
>>
>>
>> Just to stir the pot a little more:
>> 1.??? ALS has some beamlines which can go down as far as Li.? Work has been done on Li, but only on concentrated stuff.
>> 2.??? The sample is probably really dilute in Be.? The signal will be pathetic.
>> 3.??? Radiation damage will be a major issue, especially with X-ray Raman, which is a highly inefficient technique.? Even LHe cooling doesn't solve that problem.
>> 4.??? Are there model compounds available for proposed local structures of the Be center?? You'd need that to have any hope of making sense out of Be XANES.
>> 5.??? Is there such a thing as neutron PX?? That might see the Be.
>> ??? mam
>> ?
>> ----- Original Message -----
>>> From: Erik Farquhar
>>> To: XAFS Analysis using Ifeffit
>>> Sent: Tuesday, December 18, 2012 5:32 AM
>>> Subject: Re: [Ifeffit] Beryllium and EXAFS
>>>
>>>
>>> To add to Matt's last paragraph, you might also consider beamline 6-2 at SSRL (http://www-ssrl.slac.stanford.edu/beamlines/bl6-2/), as they do a fair amount of work with proteins and have an X-ray Raman setup. I think the beamline scientist occasionally?posts to this list.
>>>
>>>
>>> Good luck,
>>> Erik
>>>
>>>
>>> --
>>> Erik Farquhar, Ph.D.
>>> Case Center for Synchrotron Biosciences
>>> Brookhaven National Lab
>>> NSLS, Building 725A-X3
>>> Upton, NY? 11973
>>> efarquhar at bnl.gov
>>> +1-631-344-8174
>>>
>>>
>>>
>>> ________________________________
>>> From: Matt Newville <newville at cars.uchicago.edu>
>>> To: XAFS Analysis using Ifeffit <ifeffit at millenia.cars.aps.anl.gov>
>>> Sent: Tuesday, December 18, 2012 7:52 AM
>>> Subject: Re: [Ifeffit] Beryllium and EXAFS
>>>
>>> Hi Gina,
>>>
>>> On Mon, Dec 17, 2012 at 5:23 PM, Clayton, Gina <ClaytonG at njhealth.org> wrote:
>>>> Hi there
>>>>
>>>> I am trying to find out more about EXAFS. I am a protein crystallographer.
>>>>
>>>>
>>>> I want to identify the environment of a beryllium ion in the protein, I work with,? of which I have a crystal structure.? EXAFS was recommended as a good technique for identification of the beryllium site and species. The beryllium is an acidic pocket of the protein surrounded by oxygens from the protein, likely covalently bound. However I have had? zero result finding a beam line that can tune their setups to the Be edge at 111 eV. Also some EXAFS users say that EXAFS technique would be good for working out the environment of the Be but some say it would not . In particular? I have been advised that? Be EXAFS would not be useful since? the Be? EXAFS signal would have unacceptable interference from the carbon K-edge at 285 eV and the oxygen K edge signal at 535 eV.
>>>>
>>>> Would EXAFS work or something in a similar vein? Or, for instance,? could the acidic pocket be identified using EXAFS, by measuring the oxygens (instead of the Beryllium) using the protein with and without beryllium?
>>>>
>>>>
>>>> Thank you so much for any advice.
>>>>
>>>> Gina
>>>
>>> As you and as the person who advised you suspect, you face a number of
>>> challenges:
>>>
>>> 1.? The extremely low energy (111 eV).? You might be able to find a UV
>>> source or x-ray beamline that can reach this energy , but often such
>>> work is done in vacuo, and so may not be suitable for proteins.? Using
>>> electrons (from a TEM, say) might be possible too -- these can
>>> sometimes do C and N edges either directly or looking at Electron
>>> Energy Loss (send in an electron of high energy E1, and look for
>>> electrons of energy E1 - E, and scan the analyzer for the loss energy
>>> E through the absorption edge).? Again, electron probes may not be
>>> what you want for your proteins.
>>>
>>> 2. The C K-edge will limit your signal to "only" 175 eV.? I would say
>>> to not worry so much about this limitation.? You'll really be
>>> measuring NEXAFS (near-edge X-ray absorption ....)? but that's OK.
>>> People measure the B, C, N, and O K edges, all facing the same
>>> problem, especially for the O K edge: our planet is more or less
>>> covered with the stuff ;).? These spectra tend to be very rich and
>>> sensitive to ligands.
>>>
>>> 3. Not much (zero?? I can almost believe that!) literature on Be XAFS.
>>> This is unfortunate because most NEXAFS is measure and compare to
>>> known spectra.? I think that might mean you have to spend effort to
>>> building up a spectral library for Be in organic phases.
>>>
>>> I think trying to measure the O K edge with and without Be is nearly
>>> hopeless.? All the X-ray spectroscopies average equally over all the
>>> atoms of the absorbing species.? In addition all the spectra are
>>> grossly the same (a step function from 0 below the edge to 1 above the
>>> edge).? These make it difficult to see changes below about 1%.? ? So I
>>> think you would just have too high of an O::Be ratio to see anything
>>> useful.
>>>
>>> One option to consider is using X-ray Raman scattering
>>> (http://en.wikipedia.org/wiki/X-ray_Raman_scattering).? Here, one hits
>>> the sample with high-energy X-rays (say, 10 keV), which are highly
>>> penetrating for reasonably sized protein sample.? One analyzes an
>>> energy loss (again, E1 - E, where here E1 is around 10 keV) to get the
>>> edge of interest.? I'm familiar with (been involved with) studies on
>>> B, C, and O using this technique (all in solids, mostly at high
>>> pressure).? See, for example,? S. K. Lee, et al "Probing of bonding
>>> changes in B2O3 glasses at high pressure with inelastic X-ray
>>> scattering",? Nature Materials 4, pp 851-854 (2005), and also the work
>>> of Simo Huotari.? ? I'm not aware of anyone doing Be K-edge this way,
>>> but it should definitely be possible. For the work I've been involved
>>> with, Be would be challenging because there are other excitations in
>>> solids that appear around 100 eV.? I believe that effect would be far
>>> suppressed in a protein, though I'm not sure.
>>>
>>> That's somewhat more exotic than straightfoward XAFS,, but it might be
>>> the best chance for measuring Be in a protein.? ? I'm not sure I know
>>> any beamlines that do X-ray Raman on proteins, but I'd suggest APS
>>> beamline 20-ID or ESRF beamline as places to start.? ? The next
>>> questions would be "how many Be atoms are in your protein?", and "how
>>> much material do you have?"....
>>>
>>> Hope that helps,
>>>
>>> --Matt Newville
>>>
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>> ------------------------------
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>> Message: 2
>> Date: Tue, 18 Dec 2012 15:18:52 -0600
>> From: Matt Newville <newville at cars.uchicago.edu>
>> To: XAFS Analysis using Ifeffit <ifeffit at millenia.cars.aps.anl.gov>
>> Subject: [Ifeffit] Larch 0.9.17 available for download and testing
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>>
>> Hi Folks,
>>
>> I'm happy to announce the first full release of Larch, the next
>> generation of Ifeffit.  This includes a kit for source installation
>> and a binary installer for Windows.  A primitive Larch GUI is included
>> that provides a command-line interface and a browser / help system for
>> data in the Larch session.  Essentially all the EXAFS and XANES
>> functionality of Ifeffit 1,2 is reproduced in Larch, with several
>> significant improvements.  The current version (0.9.17) should still
>> be considered "beta" software in that bugs should be expected, but
>> Larch is now ready for detailed "field-testing" and use.  It you're
>> using Ifeffit for scripting (that is, not using the GUIs Athena,
>> Artemis, SixPack, or Prestopronto that call Ifeffit), you should start
>> using Larch now.
>>
>> Several months ago, I described the goals and status of Larch (see
>> http://millenia.cars.aps.anl.gov/pipermail/ifeffit/2012-June/010586.html),
>> which is still accurate.  Since then, the biggest effort has been in
>> documentation, finishing some of the details of Feffit() fitting, and
>> testing.
>>
>> Larch is available at
>>  http://xraypy.github.com/xraylarch/
>> and
>>  http://cars.uchicago.edu/xraylarch/
>>
>> Any feedback on any part of Larch is welcome.  I am especially
>> interested in hearing reports from people trying the Windows installer
>> -- I've tested it only on 3 Windows7 machines.  I also reiterate my
>> plea for people to get involved in the development and extension of
>> these tools.  A main motivation for Larch's implementation and design
>> was to make it very easy to add new features and experiment with new
>> algorithms and methods for analysis.  If you're interested in helping
>> Bruce or I develop these codes, please let us know.
>>
>> Some of you may have noticed that support for the Ifeffit 1.2 series
>> has been very poor recently, including the fact that building for Mac
>> OS X has fallen far behind.  With this release of Larch, I'm very
>> happy to formally end support for Ifeffit 1.2.  I'm willing to use,
>> and help others use, the older versions of Ifeffit, Athena, etc, but
>> there will be no more releases or bug fixes for the Ifeffit 1.2
>> series.  The code is available if someone else wants to maintain it.
>>
>> Thanks,
>>
>> --Matt Newville
>>
>>
>> ------------------------------
>>
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