Can I remind everyone that animals scattering is in reality *anisotropic* and therefore in some crystals, depending on the distribution of the anomalous scatterers in the protein and the symmetry of the space group, there may be relative orientations of beam and crystal in which one has lots of anomalous signal and others in which there is little. See the Templetons' work and also:
Bricogne, G., Capelli, S.C., Evans, G., Mitschler, A., Pattison, P., Roversi, P., Schiltz, M. X-ray absorption, refraction and resonant scattering tensors in selenated protein crystals: implications for data collection strategies in macromolecular crystallography J. Appl. Cryst., 38, 168-182, 2005
I suspect there have been times in which lack of signal or lots of signal have been attributed to oxidation/reduction of Se atoms, when in fact it may have been the luck of the draw of how the crystal went up into the beam.
Ciao
Pietro
Sent from my Desktop
Dr. Pietro Roversi
Oxford University Biochemistry Department - Glycobiology Division
South Parks Road
Oxford OX1 3QU England - UK
Tel. 0044 1865 275339
________________________________________
From: CCP4 bulletin board [[log in to unmask]] on behalf of James Holton [[log in to unmask]]
Sent: 07 July 2014 00:59
To: [log in to unmask]
Subject: Re: [ccp4bb] Selenomethionine crystals
I general I agree with Tim, but provided it doesn't kill your protein,
oxidation of selenomethionine can actually enhance your anomalous signal
considerably (http://dx.doi.org/10.1107/S0907444901008666).
The reason for this is because the "white line" peak in the SeMet x-ray
absorption spectrum is actually a pre-edge feature. Although the "edge"
itself represents the point when the incoming photon has just enough
energy to completely eject a core electron from the Se atom, it is also
possible for the electron to be not exactly "ejected" but merely
promoted to an empty orbital with energy a few eV below that of a free
electron in a vacuum. It is because of these extra landing sites that
the near-edge structure of the x-ray absorption specturm (XANES or
NEXAFS) can be influenced by chemistry. Specifically, the "peak" of the
normal SeMet spectrum is a 1s-4p transition
(http://dx.doi.org/10.1021/es00009a043
http://dx.doi.org/10.1107/S0909049506048898), and the more oxidized the
Se atom is, the less occupied the 4p level becomes and the bigger the
while line gets. And, of course, the bigger the while line the more f"
phasing signal you get.
Unfortunately, the extra oxygens stuck on the Se can sterically disrupt
the local environment of the SeMet, and fear of this is probably why so
many people have not tried it. However, if you're desperate for that
extra little "boost" of phasing signal, it might be a refreshing change
to do the opposite of trying to keep your protein from oxidizing all the
time. Perhaps even adding a dash of H2O2. The worst your crystals can
do is not diffract.
-James Holton
MAD Scientist
On 7/1/2014 8:54 AM, Tim Gruene wrote:
> Dear Maher,
>
> as far as I understand, the anomalous scattering comes from inner shell
> electrons, not the valence electrons. So while you might notice a slight shift
> in the peak wavelength, the strength of the signal will only reduce if you
> crystal order suffered over time, but not from any oxidation of the Se.
>
> Best,
> Tim
>
> On Tue, Jul 01, 2014 at 10:51:20AM -0500, Maher Alayyoubi wrote:
>> Hi everyone,
>>
>> Would anyone know for how long Selenomethionine derivative crystals are
>> good if kept in plate at RT. In other words, would SE loose its scattering
>> properties due to oxidation over time? I have SElmet crystals that have
>> been lying in a plate for 2 months by now so I was wondering if they are
>> still worthwhile the effort of collecting new data from them.
>>
>> Thank you very much,
>>
>>
>> Maher
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