Crystallographic Methods and Protocols / Methods in Molecular Biology Bd.56 (PDF)

The volumes in the series, Methods in Molecular Biology, are conceived with the biochemist and molecular biologist in mind. The present book, Crystallographic Methods andProtocols, concentrates on the use of X-ray crystallography to solve the detailed three­ dimensional structuresofproteins, nucleic acids, andtheir complexes. Such a structure determination is a major undertaking, demands expertise in a range of skills, and requires considerable resources. The biologically trained worker will probably first become involved when identifying an important scientific problem whose solution would benefit from a full structure. The protein or nucleic acid at issue must be sequenced and prepared to high purity in appropriate quantities, probably by either chemical synthesis for nucleic acids or genetic engineering for proteins. CrystallographicMethods andPro­ tocols aims to give biologically trained workers an insight into the techniques used to crystallize their proteins, obtain the raw X-ray data, and solve and refine the structure. The aim ofa crystal structure determination is to provide infor­ mation that will solve biologically relevant problems; that process normally requires a high resolution structure. The preparationofsuit­ able crystals for a high resolution structure remains the major bottle­ neck in structure determination, and the effective application of appropriate genetic engineering and biochemical techniques in the initial stages pays ahandsome dividend later. The productionofsuit­ able proteins by recombinant methods and the preparationofcrystal­ line derivatives are covered by Skelly and Madden in Chapter 2. Before beginning the data collection required for a high resolution structure, a preliminary characterizationofthe crystals is carried out.