The overall objective of the proposed research is to make the power of anomalous-scattering methods effective in the full range of biological crystallography. The plan builds on accomplishments of this project and worldwide experience, showing that multiwavelength anomalous diffraction (MAD) is now the mainstay of de novo structure determination. We propose to explore the optimization of MAD and SAD, (MAD's single-wavelength counterpart), for applications ranging from structural genomics to membrane proteins and macromolecular assemblages and on to lead discovery for drug development. Realization of the full potential requires further development in instrumentation, in computational and experimental procedures, and in biochemistry. Our anomalous-diffraction experiments will be conducted primarily at synchrotron beamlines that we have constructed at Brookhaven National Laboratory. The overall objective is embodied in four specific aims: (1) We propose to enhance the theoretical and computational tools for analyzing anomalous-diffraction experiments. (2) We propose to develop improved and more conveniently accessible instrumentation and more effective experimental procedures for measuring anomalous diffraction. (3) We propose to develop alternative means for introducing anomalous-scattering centers into macromolecules and to characterize the resonant features associated with these labels. (4) We propose to advance the development of phase evaluation from anomalous-scattering measurements through MAD and SAD applications to challenging problems of biological and medical significance. Biologically exciting problems motivate the development of appropriate tools, and forefront methods to accelerate the solution of interesting structures. ? ?
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