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Radiation damage affects MAD experiments in two ways: (i) increased absorption by the crystal at the wavelengths of interest for the experiment results in faster crystal deterioration; (ii) lack of isomorphism induced by radiation damage causes problems when scaling and merging data at different wavelengths and can prevent accurate measurement of anomalous and dispersive differences. In an attempt to overcome these problems in the case of radiation-sensitive crystals of vinculin, two-wavelength MAD data were collected at the Se absorption-edge inflection and at high-energy remote wavelengths. Although this strategy resulted in a lower total absorbed dose compared with a standard three-wavelength experiment using the peak wavelength, an increase in the unit-cell volume and other effects attributable to radiation damage were still observed. In an effort to extract the maximum information available from the data, different data-processing and scaling procedures were compared. Scaling approaches involving local scaling of unmerged reflections were consistently successful and most ordered Se sites could be located. Subsequent use of these sites for phasing resulted in an interpretable electron density map. This case demonstrates the feasibility of two-wavelength MAD in the presence of moderate radiation damage using conventional data collection strategies and widely available standard software.

Original publication

DOI

10.1107/S0909049505003249

Type

Journal article

Journal

J Synchrotron Radiat

Publication Date

05/2005

Volume

12

Pages

285 - 291

Keywords

Crystallization, Crystallography, X-Ray, Feasibility Studies, Protein Conformation, Software, Vinculin