Can we find solution?

1. Fitting EM maps into X-ray Data Alexei VaginYork Structural Biology LaboratoryUniversity of York

2. Can we find solution? Weight of low resolution grows Low high Model Similarity 0 .1 0.2 0.3 0.4 0.5 Very likely Weight of high resolution grows Low high Model size 0.5 0.4 0.3 0.2 0.1 Very unlikely EM model

3. Information in X-ray and EMmust overlap Resmin X-ray s Resmax EM

4. Example 1 Groel-ATP7-Groes Map: EMD-1180 Fitted atomic model: PDB 2c7c X-ray: PDB 1sx4 ( 40 - 3 A )

5. EM map as model

6. EM map (sfcheck) from grid

7. EM map (sfcheck) 7A

8. Information in X-ray and EMmust overlap Resmin X-ray (40A) s Resmax EM (7A)

9. ∆Ph =⃒Phxray - Phem⃒ MR solution (EM model)∆Ph =⃒Phxray - Phmodel⃒MR solution (fitted atomic model) ∆Ph 90o 50o Res: 11.1 9.1 7.7 6.7 5.9 5.3 4.8 4.3 4.0 A

10. Map ( Fobs Phem ) 6Å

11. What we can do with these phases 1. Help to find HA positions in the derivative 2. Extend phases by averaging electron density 3. Try to fit some fragments ( helix )

12. EM map (4A) Modified map Averaging and Solvent flattening

13. Map (FobsPhem) 4Å Modified map

14. ∆Ph =⃒Phxray - Phem⃒ MR solution (EM model)∆Ph =⃒Phxray - Phmodel⃒ MR solution (fitted atomic model)∆Ph =⃒Phxray - Phaver⃒MR solution (after DM) ∆Ph 90o 50o Res: 11.1 9.1 7.7 6.7 5.9 5.3 4.8 4.3 4.0 A

15. Free atom model

16. ∆Ph =⃒Phxray - Phem⃒ MR solution (EM model)∆Ph =⃒Phxray - Phmodel⃒ MR solution (fitted atomic model)∆Ph =⃒Phxray - Phmodel_ref⃒ MR solution (refined atomic model)∆Ph =⃒Phxray - Phfree⃒ MR solution (free atom model after DM) ∆Ph 90o 50o Res: 9.4 7.4 6.1 5.2 4.5 4.0 3.6 3.3 3.0 A

17. Map (FobsPhem) 3Å Map (FobsPhfree) 3Å (Free atom model after DM)

18. Map (FobsPhfree) 3Å (Free atom model after DM)

19. Map (FobsPhfree) 3Å (Free atom model after DM)

20. Example 2 X-ray structure of GP6 protein of phage SPP1 ( 3.4 A ) Fred Antson at al. University of York

21. EM map as model

22. EM map (sfcheck 1) from grid

23. EM map (sfcheck 2) 8A

24. Information in X-ray and EMmust overlap Resmin X-ray (40A) s Resmax EM (8A)

25. ∆Ph =⃒Phxray - Phem⃒ MR solution (EM model) ∆Ph 90o 50o Res: 10.3 8.3 6.9 6.0 5.2 4.7 4.2 3.8 3.5 A

26. Map ( Fobs Phem ) 7Å

27. What we can do with these phases 1. Help to find HA positions in the derivative 2. Extend phases by averaging electron density 3. Try to fit some fragments ( helix )

28. ∆ Ph =⃒Phxray - Phem⃒ MR solution (EM model)∆ Ph =⃒Phxray - Phaver⃒ MR solution (after DM) ∆Ph 90o 50o Res: 10.3 8.3 6.9 6.0 5.2 4.7 4.2 3.8 3.5 A

29. Free atom modelmodel was used only to compute initial phases

30. ∆ Ph =⃒Phxray - Phem⃒ MR solution (EM model)∆ Ph =⃒Phxray - Phaver⃒ MR solution (after DM)∆ Ph =⃒Phxray - Phfree⃒ MR solution (free atom model after DM) ∆Ph 90o 50o Res: 10.3 8.3 6.9 6.0 5.2 4.7 4.2 3.8 3.5 A

31. Final map 3.5 A

32. Final map 3.5 A

33. Map ( Fobs Phem ) Final map

34. What we can do with these phases 1. Help to find HA positions in the derivative 2. Extend phases by averaging electron density 3. Try to fit some fragments ( helix )

35. Fitting 17 x 13 helixes into final map by SAPTF

36. Final map 3.5 A and helixes

37. X-ray model and helixes

38. Model: helix_17x13.pdb 1.Refinement 2.Compute map for refined model 3.Combine final map and map from model Final map Combine map

39. ∆ Ph =⃒Phxray - Phem⃒ MR solution (EM model)∆ Ph =⃒Phxray - Phaver⃒ MR solution (after DM)∆ Ph =⃒Phxray - Phfree⃒ MR solution (free atom model after DM)∆ Ph =⃒Phxray - Phcomb⃒ combine map ∆Ph 90o 50o Res: 10.3 8.3 6.9 6.0 5.2 4.7 4.2 3.8 3.5 A

40. The end http://www.ysbl.york.ac.uk/~alexei/

41. SOLUTION_CHECK:Are twosolutions identical?Alexei VaginYork Structural Biology LaboratoryUniversity of York

42. It is trivial. Xtest [Torigin] [Tsymm_op] Ytest Xtarget Alignment, Fitting Orientation and position differences

43. The end