Fitting EM maps into X-ray Data Alexei Vagin York Structural Biology Laboratory University of York

EM  X-ray ( P2 ) --> MR --> H2OH2O H2OH2O

Molecular Replacement Cell Model 1. Rotation function 2.Translation function

Can we find solution? Weight of low resolution grows Low high Model Similarity Weight of high resolution grows Low high Model size Very likely Very unlikely EM model

Information in X-ray and EM must overlap s Resmin X-ray Resmax EM

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

EM map as model

EM map (sfcheck) from grid

EM map (sfcheck) 7A

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

∆Ph =⃒Ph xray - Ph em ⃒ MR solution (EM model) ∆Ph =⃒Ph xray - Ph model ⃒ MR solution (fitted atomic model) 50 o 90 o Res: A ∆Ph

Map ( F obs Ph em ) 6Å

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 )

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

Map (F obs Ph em ) 4Å Modified map

∆Ph =⃒Ph xray - Ph em ⃒ MR solution (EM model) ∆Ph =⃒Ph xray - Ph model ⃒ MR solution (fitted atomic model) ∆Ph =⃒Ph xray - Ph aver ⃒ MR solution (after DM) 50 o 90 o Res: A ∆Ph

Free atom model

∆Ph =⃒Ph xray - Ph em ⃒ MR solution (EM model) ∆Ph =⃒Ph xray - Ph model ⃒ MR solution (fitted atomic model) ∆Ph =⃒Ph xray - Ph model_ref ⃒ MR solution (refined atomic model) ∆Ph =⃒Ph xray - Ph free ⃒ MR solution (free atom model after DM) 50 o 90 o Res: A ∆Ph

Map (F obs Ph em ) 3Å Map (F obs Ph free ) 3Å (Free atom model after DM)

Map (F obs Ph free ) 3Å (Free atom model after DM)

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

EM map as model

EM map (sfcheck 1) from grid

EM map (sfcheck 2) 8A

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

∆Ph =⃒Ph xray - Ph em ⃒ MR solution (EM model) 50 o 90 o Res: A ∆Ph

Map ( F obs Ph em ) 7Å

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 )

∆ Ph =⃒Ph xray - Ph em ⃒ MR solution (EM model) ∆ Ph =⃒Ph xray - Ph aver ⃒ MR solution (after DM) 50 o 90 o Res: A ∆Ph

Free atom model model was used only to compute initial phases

∆ Ph =⃒Ph xray - Ph em ⃒ MR solution (EM model) ∆ Ph =⃒Ph xray - Ph aver ⃒ MR solution (after DM) ∆ Ph =⃒Ph xray - Ph free ⃒ MR solution (free atom model after DM) 50 o 90 o Res: A ∆Ph

Final map 3.5 A

Map ( F obs Ph em ) Final map

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 )

Fitting 17 x 13 helixes into final map by SAPTF

Final map 3.5 A and helixes

X-ray model and helixes

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

∆ Ph =⃒Ph xray - Ph em ⃒ MR solution (EM model) ∆ Ph =⃒Ph xray - Ph aver ⃒ MR solution (after DM) ∆ Ph =⃒Ph xray - Ph free ⃒ MR solution (free atom model after DM) ∆ Ph =⃒Ph xray - Ph comb ⃒ combine map 50 o 90 o Res: A ∆Ph

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SOLUTION_CHECK: Are two solutions identical? Alexei Vagin York Structural Biology Laboratory University of York

It is trivial. X test [T origin ] [T symm_op ] Y test X target Alignment, Fitting Orientation and position differences

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