Theoretical Aspects

Summary

How to use MODULE
 

Required Material

Starting the Program

Input File Format

Initial Display

Displaying the Primary Sequence

Selection of Type of Coupling

Setting Internuclear Distances

Fitting Single Modules

Fitting the Alignment Tensor

Correlation Plots

Comparision of Calculated and Experimental Couplings

Monte Carlo Error Analysis

Test Sample 1

Choice of Modules from Primary Sequence

Fitting the Alignment Tensors

Correlation Plots

Comparision of Calculated and Experimental Couplings

Monte Carlo Error Analysis

Common Alignment Frame

Degenerate Orientations

Automatic Calculation of Molecular Architecture

Covalent Bonds

Test Sample 2

Special Cases

Axially Symmetric Alignment Tensor.

Highly Rhombic Alignment Tensor.

 
Distance Constraints

Creating and Reading Work Folders

Simulating Datasets

Hammerhead Ribozyme - Orientation of Secondary Structural Motifs.

Test Sample 3

Protein-Protein Complexes.

Test Sample 4
 
 

Fitting the Alignment Tensor

To fit the tensor to the selected couplings with this parameter set simply click on Fit in the main MODULE window - This should take a few seconds on a R10000 SGI processor (approximately equivalent to a 400MHz PC)
 
 

Having fitted the tensor, the axial and radial components (A_a and A_r) and the euler angles describing the orientation of the tensor relative to the pdb frame, are shown in the background window - NOTE that the axial and rhombic components are quoted and treated in absolute order units throughout the program (normally 10^-4 is cited).

The tensor can then be visualised with respect to the pdb coordinates (Orientation in the Visualisation menu) -
 
 

Correlation Plots

To check the quality of the fit, or to identify outliers, correlations of calculated and experimental couplings can be viewed, either separately or for all couplings together by selecting the option Back-Corr in the Visualisation menu:

Selecting Separate Couplings gives the correlations between calculated and experimental for each different type of coupling, defined by the different atom types -
 
 

The correlation for each particular coupling type can be viewed in detail using the same menu. The cursor can be used to identify the specific couplings (for example the outlier shown here).

In example shown below we have selected the CO-N coupling :
 
 

Note that in each of these options the local chi2 is shown (in blue at the bottom of the plot), with respect to the individual points. This allows outliers or problematic couplings to be identified easily. A text line in the window (at the top) gives the name, chi2 and calculated and experimental values of the coupling corresponding to the cursor poisition (yellow line).
 
 
 

Comparision of Calculated and Experimental Couplings
 

Similarly, the experimental and calculated couplings can be visualised with respect to the primary sequence, either separately or for all couplings together by selecting the option Back-Diff in the Visualisation menu:
 
 

Separate Couplings -
 
 

HN-CO Couplings

Monte Carlo Error Analysis

The uncertainty associated with the orientation of the tensor axes, and the values of the axial and rhombic components can be estimated using a Monte-Carlo based error analysis.

This analysis takes the best-fit tensor and back-calculates simulated datsets from this using a Gaussian noise distribution.

The noise distribution is based on the data uncertainty in the initial input file.

The Monte Carlo analysis can be selected using the button in the main display window.
 
 

The angular dispersion is small in this case - the noise which was simulated for this 'Dataset' was based on an uncertainty of 5% so this is perhaps not surprising.

The dispersion in the magnituide of Da and Dr can be visualised using the Monte Carlo in the Visualisation menu
 
 

In the example shown below the tensor has axial symmetry, so the transverse components of the tensor are equivalent (Axx-Ayy=0). The results of the monte-carlo simulation reflect this; the dispersion in the transverse plane is continuous, while the direction of the Azz component is well defined. Note that there are points in both positive and negative z-directions.
 
 

Test Sample 1 - The files used for this example are enclosed with the downloaded package and are called -
 
 

sample1.pdb

sample1.dat