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        Use of Multiple-Wavelength Anomalous Diffraction Measurements in Ab Initio Phase Determination for Macromolecular Structures

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        The determination of three-dimensional structures of macromolecules in the crystalline state depends on the acquisition and processing of diffraction data generated by the interaction of X-rays with the crystals of the macromolecule under investigation. In general, there are two points at which the process languishes because of lack of necessary material or data. The first is the initial crystallization of the macromolecule; although work is being done to understand the principles involved (1 ), the crystallization of a macromolecule usually depends heavily on empirical reasoning (2 ). The second bottleneck is acquisition of phases. Each diffraction maximum has associated with it both an amplitude and a phase. The phases are not directly measurable in a typical diffraction experiment and must therefore be estimated by indirect means. By far the most widely used method for the derivation of phases is that of Multiple Isomorphous Replacement (MIR). The MIR method depends on the making of isomorphous derivative crystals of the macromolecule under investigation, and estimation of phases from the differences between the amplitudes of native and derivative diffraction maxima (3 ). At least two isomorphous derivatives are required for reliable phase determination, although in favorable cases, anomalous diffraction information from a derivative may be used as a second pseudoderivative.
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