2VW7

Nitrite reductase from Alcaligenes xylosoxidans - 1 of 3

Summary for 2VW7

• Entry DOI: 10.2210/pdb2vw7/pdb
• Related: 1BQ5 1GS6 1GS7 1GS8 1HAU 1HAW 1NDT 1OE1 1OE2 1OE3 1WA0 1WA1 1WA2 1WAE 2BO0 2BP0 2BP8 2JFC 2VM3 2VM4 2VMJ 2VN3 2VW4 2VW6
• Descriptor: DISSIMILATORY COPPER-CONTAINING NITRITE REDUCTASE, COPPER (II) ION, ZINC ION, ... (5 entities in total)
• Functional Keywords: nitrite reductase, denitrification, microspectrophotometer, ordered mechanism, oxidoreductase
• Biological source: ACHROMOBACTER XYLOSOXIDANS
• Total number of polymer chains: 2
• Total formula weight: 73914.51

Authors

Ellis, M.J.,Buffey, S.G.,Hough, M.A.,Hasnain, S.S. (deposition date: 2008-06-16, release date: 2008-09-09, Last modification date: 2024-05-01)

Primary citation

Ellis, M.J.,Buffey, S.G.,Hough, M.A.,Hasnain, S.S.
On-Line Optical and X-Ray Spectroscopies with Crystallography: An Integrated Approach for Determining Metalloprotein Structures in Functionally Well Defined States.
J.Synchrotron Radiat., 15:433-, 2008

PubMed Abstract: X-ray-induced redox changes can lead to incorrect assignments of the functional states of metals in metalloprotein crystals. The need for on-line monitoring of the status of metal ions (and other chromophores) during protein crystallography experiments is of growing importance with the use of intense synchrotron X-ray beams. Significant efforts are therefore being made worldwide to combine different spectroscopies in parallel with X-ray crystallographic data collection. Here the implementation and utilization of optical and X-ray absorption spectroscopies on the modern macromolecular crystallography (MX) beamline 10, at the SRS, Daresbury Laboratory, is described. This beamline is equipped with a dedicated monolithic energy-dispersive X-ray fluorescence detector, allowing X-ray absorption spectroscopy (XAS) measurements to be made in situ on the same crystal used to record the diffraction data. In addition, an optical microspectrophotometer has been incorporated on the beamline, thus facilitating combined MX, XAS and optical spectroscopic measurements. By uniting these techniques it is also possible to monitor the status of optically active and optically silent metal centres present in a crystal at the same time. This unique capability has been applied to observe the results of crystallographic data collection on crystals of nitrite reductase from Alcaligenes xylosoxidans, which contains both type-1 and type-2 Cu centres. It is found that the type-1 Cu centre photoreduces quickly, resulting in the loss of the 595 nm peak in the optical spectrum, while the type-2 Cu centre remains in the oxidized state over a much longer time period, for which independent confirmation is provided by XAS data as this centre has an optical spectrum which is barely detectable using microspectrophotometry. This example clearly demonstrates the importance of using two on-line methods, spectroscopy and XAS, for identifying well defined redox states of metalloproteins during crystallographic data collection.

PubMed: 18728313
DOI: 10.1107/S0909049508014945

Experimental method

X-RAY DIFFRACTION (1.9 Å)