We have facilities for using xenon derivatives (at room temp. and 100K) for
MIR and Anomalous Dispersion experiments on the protein crystallography stations
at the SRS, Daresbury.
Users of the xenon pressure cell at Daresbury are required to complete a
class 3 risk assessment, including a description of the hazard and safe system
of work, prior to their beamtime.
http://www.nwsgc.ac.uk/beamline10/xenon/new_risk.html
Why Use XENON?
- Derivative protein crystals can be produced by pressurising native crystals with xenon gas. Modification of the mother liquor to determine soaking conditions is avoided.
- The number of binding sites, and their occupancies can be changed by altering the gas pressure, thus, several derivatives could be produced from the same crystal. These sites often differ from metal binding sites.
- Xenon atoms interact weakly with protein; isomorphism of the derivative with the native is high.
- Xenon binding is often reversible; the same crystal could be used for a further heavy atom soak.
Problems
- Systematic errors in measured intensities due to absorption by the pressurising gas.
- Accelerated radiation damage to unfrozen crystals.
- Safety implications associated with high pressure equipment.
- Possible formation of hydrates at low temperatures and high pressures.
XENON Properties
| XENON (Xe) | |
|---|---|
| Atomic no | 54 |
| Melting point | -112 |
| Boiling point | -108 |
| Relative density (Gas) | 4.5 |
| Relative density (liquid) | 1.5 |
| Molecular weight | 131 |
| Solubility in water | 644 mg/l |
| Absorption edges (Å and (keV)) | ||||
|---|---|---|---|---|
| K | LI | LII | LIII | MI |
| 0.3587 | 2.2738 | 2.4293 | 2.5926 | 10.8321 |
| (34.5614) | (5.4528) | (5.1037) | (4.7822) | (1.1446) |
Text file of anomalous
scattering coefficients f' and f'' as a Function of energy
For Frozen Crystals
We have acquired an Oxford Cryosystems Xcell for pressurising crystals prior to freezing.
Users of the xenon pressure cell at Daresbury are required to complete a
class 3 risk assessment, including a description of the hazard and safe system
of work, prior to their beamtime.
http://www.nwsgc.ac.uk/beamline10/xenon/new_risk.html
Instructions for Using the Oxford Cryosystems Xcell
-
Unscrew the Xcell filter paper cap and replace the old filter with one soaked in mother liquor. Refit the filter paper cap and tighten finger tight. Do not remove the filter paper cap once the Xcell is pressurised.
-
Unlock the Xcell slider mechanism and fit a magnetic base to the sample mount.
-
Connect the gas regulator to the xenon gas bottle.
-
Connect the gas hose to the regulator and the Xcell inlet valve. Do not disconnect the gas hose from the regulator once the hose is pressurised. Do not use the Xcell on uneven surfaces.
-
Ensure: the Xcell vent valve is closed; the slider mechanism is locked; the filter paper cap is fitted and the inlet valve is closed.
-
Vent the Xcell and gas hose by: opening the xenon gas bottle valve; setting the gas regulator to the working pressure; slowly opening the Xcell inlet valve; slowly opening the Xcell vent valve to bleed air from the system. Do not exceed 25 bar when pressurising the Xcell
-
Close the Xcell inlet and vent valve.
-
Disconnect the Xcell from the gas hose and position it close to the sample. Do not use the Xcell on uneven surfaces.
-
Slowly open the Xcell vent valve to de-pressurise the chamber.
-
Unlock the Xcell slider mechanism and expose the magnetic base.Do not open the chamber while the Xcell is pressurised.
-
Mount the loop with crystal on the magnetic base.
-
Shut and lock the Xcell slider mechanism.
-
Carefully take the Xcell back to the xenon gas bottle.
-
Reconnect the Xcell to the gas hose. Do not use the Xcell on uneven surfaces.
-
Ensure: the Xcell vent valve is closed; the slider mechanism is locked; the filter paper cap is finger tight and the inlet valve is closed.
-
Slowly open the Xcell inlet valve pressurising the chamber to the working pressure. Do not exceed 25 bar.
-
Slowly bleed air from the Xcell chamber using the vent valve.
-
Ensure the Xcell has reached the working pressure then close the inlet vale. Do not exceed 25 bar.
-
Close the xenon gas bottle valve.
-
Leave the crystal under pressure for the desired length of time.
-
Disconnect the Xcell from the gas hose.
-
Carefully position the Xcell close to the freezing agent. Do not use the Xcell on uneven surfaces.
-
Slowly open the Xcell vent valve to de-pressurise the chamber.
-
Unlock the slider mechanism, carefully remove the loop and crystal for freezing. Do not open the chamber while the Xcell is pressurised.
Future Developments
Explore the possibilities of exploiting absorption edges for MAD and SAD experiments. Xe K-edge 0.3587 Å, Kr K-edge 0.86552 Å.REFERENCES
- Structure of lobster apocrustacyanin A1 using softer X-rays. M. Cianci, P.
J. Rizkallah, A. Olczak, J. Raftery,
N. E. Chayen, P. F. Zagalsky
and J. R. Helliwell (2001). Acta
Cryst. D57, 1219-1229.
- Better structures from better data through better methods: a review of
developments in de novo macromolecular phasing techniques and associated
instrumentation at LURE. Fourme, R. et al (1999) J. Synch. Rad. 6(4)
834-844
- A method to stabilize reduced and/or gas treated protein crystals by flash
cooling under a controlled atmospher
Xavier Vermede et.al. J. App. Cryst, (1999) 32(3) 505-509
- Protein Crystallography at Ultra-Short Wavelengths: Feasibility Study of
Anomalous Dispersion Experiments at the Xenon K-Edge. Schiltz, M., Kvick,
A., Svensson, O., Shepard, W., De LaFortelle, E., Prange, T., Kahn, R. &
Fourme, R. (1997). Journal of Synchrotron Radiation , 4,
287-297.
- Freeze-Trapping Isomorphous Xenon Derivatives of Protein Crystals. Sauer,
O.; Schmidt, A.; Kratky, C. (1997). Journal of Applied Crystallography,
30, 476-486.
- Successful flash-cooling of xenon-derivatised myoglobin crystals. Soltis,
S.M.; Stowell, M.H.B.; Wiener, M.C.; Philips, G.N.; Rees, D.C. (1997).Journal
of Applied Crystallography, 30, 190-194.
- On the Preparation and X-ray Data Collection of Isomorphous Xenon
Derivatives. Schiltz, M.; Prange, T.; Fourme, R. (1994). Journal of
Applied Crystallography, 27, 950-960.
- Using Xenon as a Heavy Atom for Determining Phases in Sperm Whale
Metmyoglobin. Vitali, J.; Robbins, A.H.; Almo, S.C.; Tilton, R.F. (1991). Journal
of Applied Crystallography, 24, 931-935.
- Cavities in proteins: structure of a metmyoglobin-xenon complex solved to
1.9&Angring. Tilton, R.F.; Kuntz, L.D.; Pesko, G.A. (1984) Biochemistry
23. 2849-2857.
- Binding of Xenon to Sperm Whale Myoglobin. Schoenborn B.P.; Watson, H.C.;
Kendrew, J.C. (1965). Nature, 207, 28-30.
