Comparing Data from MD simulations and X-ray Crystallography What can we compare? 3D shapes (Scalar coupling constants, a.k.a. J-values, nuclear Overhauser effects) Atomic motions (RMSF, Order Parameters)

NOE intensities may be used to estimate the distance between hydrogen atoms, separated by less than 4 Å J-couplings may be used to estimate the torsion angles between the rings Sparse, complex data!

NMR structure of CCHF virus Gn tail zinc finger. Estrada D F, De Guzman R N J. Biol. Chem. 2011;286: ©2011 by American Society for Biochemistry and Molecular Biology NMR structure of CCHF virus Gn tail zinc finger. Stereoview of the superposition of 20 lowest energy NMR structures of CCHF virus Gn zinc finger (A).

NMR Scalar Couplings (J-values) Depend on Torsion Angles NMR – measure J-value from spectrum then derive angle MD – compute angle from trajectory then derive J-value

Nuclear Overhauser Effects The nuclear Overhauser effect (NOE) identifies pairs of protons (i,j) that are in close proximity. (A) Schematic representation of a polypeptide chain highlighting five particular protons. Protons 2 and 5 are in close proximity Under certain NMR conditions: the magnitude of the NOE intensity is proportional to 1/R ij 6, where Rij is the distance between the protons. Therefore, NOEs can also be computed from MD data (by computing 1/R ij 6 ) (A) NOESY observations show that protons (connected by dotted red lines) are close to one another in space. (B) A three-dimensional structure calculated with these proton pairs constrained to be close together i j R ij

NMR Order Parameters Relate to Internal Motion Elapsed Time   Initially the structures will align closely (highly ordered) and the angle between any internal vector will be close to zero but later the structures will be very different (disordered). Order Parameter versus Simulation time

For example – when comparing to x-ray data, it is possible to compare theoretical atomic positions or interatomic distances with experimental values – but remember the x-ray data were not measured under the same conditions as the simulation – when comparing to NMR data, remember, 3D structures reported from NMR studies are all models based on fitting to the measured NMR observables (NOE intensities and couplings). It may be preferable to compute these NMR observables from the theoretical data, rather than comparing to a 3D model derived from the NMR data

PropertyX-RayNMRMD Simulation Atomic PositionsWell DefinedNot DefinedWell Defined Bond Lengths and Angles Well DefinedNot DefinedWell Defined Inter Atomic Distances Well DefinedPartially Defined by NOEs Well Defined Torsion AnglesWell DefinedPartially Defined by J-values Well Defined Internal MotionsNot DefinedPartially Defined by correlation times Well Defined

P 2 (x) = ½(3x 2 - 1) a. b = |a||b|cos  1) Run the MD 2) monitor the N-H bond vectors 3) compute C(t) 4) Derive S 2 5) Compare to S 2 from NMR 1)Run the MD 2)Monitor the dihedral angles 3)Compute scalar J-values 4)Derive a simple relationship to assess the goodness of fit, X 2 5)Compare to NMR

Perfect agreement would give X 2 = 0 So which water model is superior? Why did they increase the system temperature from 300 to 350 K?

Is a X 2 value of 3Hz a big error? Based on the graph below, what range of torsion values is defined by 3Hz?

Vary the force field parameters and see what set gives best agreement with NMR But how do you know what parameter to vary?

Why does the C-terminus of the protein have a low value for the order parameter? Because it is highly flexible…