From Time-Averaged to Time-Resolved Crystallography: Studies on Superoxide Dismutase and Myoglobin

Research output: ThesisDoctoral Thesis (compilation)


We have studied protein motions with nanosecond time-resolved crystallography. The release of carbon monoxide from its complex with myoglobin (MbCO) was triggered by nanosecond laser pulses. X-ray data were collected with time delays ranging from 4 ns to 1.9 ms using the Laue method, at the European Synchrotron Radiation Facility in Grenoble. For the first time point, X-ray pulses of 150 ps duration were used. Electron difference maps show the release of the CO and the subsequent motion of the iron out of the haem plane. In the 4 ns difference map, a positive density feature is found at a position coinciding with that of an intermediate docking site seen at low temperature. The following time points show the rebinding of the CO, and also indications of protein relaxations extending further than 4 ns. This is the first time-resolved protein crystallography experiment with nanosecond time-resolution yielding structural results.

We have developed instrumentation and methods for time-resolved protein crystallography, including a method to reduce the noise in electron density difference maps from poorly accurate data. The method is also applicable to other cases where the signal to noise ratio is low.

The crystal structure of iron superoxide dismutase (SOD) from the hyperthermophilic archaeon <i>Sulfolobus solfataricus</i> has been determined to 2.3 Å resolution by molecular replacement. The structure revealed an increased number of inter-subunit ion-pairs in a compact tetramer. We suggest this to be important for the thermostability. However, the general fold is found to be similar to other known iron- or manganese SOD structures. In addition, the electron density maps revealed an unexpected and unusual covalent modification of a conserved tyrosine in the active site.
Original languageEnglish
Awarding Institution
  • Biochemistry and Structural Biology
  • [unknown], [unknown], Supervisor, External person
Award date1998 Sept 26
ISBN (Print)91-628-3116-X
Publication statusPublished - 1998

Bibliographical note

Defence details

Date: 1998-09-26
Time: 10:15
Place: Lecture hall C, Chemical Center, Sölvegatan 39, Lund

External reviewer(s)

Name: Hajdu, Janos
Title: Prof
Affiliation: Uppsala university


Article: Common Features Involved in Protein Hyperthermostability derived from the Crystal Structure of Iron Superoxide Dismutase from the Archaeon Sulfolobus solfataricusUrsby, T., Adinolfi, B.S., Al-Karadaghi, S., De Vendittis, E. & Bocchini, V. (1998)Submitted to Journal of Molecular Biology.

Article: New Opportunities for Time Resolved X-ray Scattering at the ESRFWulff, M., Ursby T., Bourgeois, D., Schotte, F., Zontone, F. & Lorenzen, M. (1996)Journal de Chimie Physique et de Physico-Chimie Biologique, 93, 1915-1937.

Article: Feasibility and Realization of Single-Pulse Laue Diffraction on Macromolecular Crystals at ESRFBourgeois, D., Ursby, T., Wulff, M., Pradervand, C., LeGrand, A., Schildkamp, W., Labouré, S., Srajer, V., Teng, T.-Y., Roth, M. & Moffat K. (1996).Journal of Synchrotron Radiation, 3, 65-74.Improved Estimation of Structure Factor Difference Amplitudes from Poorly Accurate DataUrsby, T. & Bourgeois, D. (1997)Acta Crystallographica, A53, 564-575.

Article: Photolysis of the Carbon Monoxide Complex of Myoglobin: Nanosecond Time-Resolved CrystallographySrajer, V., Teng, T.-Y., Ursby, T., Pradervand, C., Ren Z., Adachi, S., Schildkamp, W., Bourgeois, D., Wulff, M. & Moffat K. (1996)Science, 274, 1726-1729.

Subject classification (UKÄ)

  • Biological Sciences

Free keywords

  • superoxide dismutase
  • Bayesian statistics
  • electron difference maps
  • synchrotron
  • docking site
  • photolysis
  • myoglobin
  • Time-resolved crystallography
  • Laue
  • thermostability
  • Sulfolobus solfataricus
  • Molecular biophysics
  • Molekylär biofysik


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