Bioc585 2008 Exercise 4 – MAD Phased Model Answers (10 points) (1) Starting with the warpNtrace map and model, build in side chains for Gly 11 T, Gly 12 T and Gly 13 T. Refine this model with REFMAC and examine the new model and map. If this model looks good (matches the density well), copy and paste an image into your answer. If not, pick a different residue and try again (hint – watch Gly 12 T carefully). Also copy and paste an image of your refinement log graphs for R-factor and geometry (the last graph, Rfactor analysis, stats vs cycle). The model builds well as Ser-Leu-Leu (density on left). (2) View the density for these three residues in the beginning experimental map (using solve.mtz). How do they compare? The density in the refined warpNtrace map is better, indicating the density modification and model building were improving the phases. The density for Ser 11 is poor (density on right, above), but that for Leu 12 and Lue 13 is reasonably convincing.(3) Examine the positions for the selenium atoms (from crystal 1, see introduction) and compare with the warpNtrace model (based on crystal 2). How well do they align with the methionine sulfurs? Which methionine is associated with which selenium? Suggest an explanation for any deviations. Se 458 corresponds to Met 54 and the two atoms are close but not exactly aligned. Se 459 corresponds to Met 42 and Se 460 to Met 44. Again, the SE atoms are close but not exactly in the same place as the sulfur atoms. The small deviations probably have to do with small differences in the unit cells of the two proteins, which differ by a few tenths of an angstrom. Se 458: Met 54, good agreement Se 459: Met 42, good agreement, different protein molecule Se 460: Met 44, good agreement, same protein molecule as Met 42 (4) Are the selenium atoms all attached to the same molecule in the unit cell? Please explain your conclusion. No. Se 458 is attached to a different protein molecule in a different asymmetric unit than are Se 459 and Se 460. The two molecules are related by crystallographic
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