In this Tutorial you learn how to make and visualize conformational trajectories based on PCA. The method is also described in Haselbach, Schrader et al.. Apart from basic functionalities of the Cow this tutorial requires the use of flow control elements.

Input for the analysis should be several 3D Volumes, that have the same size and are aligned on each other. To gain these we recomment the use of relion in combination with UCSF Chimera. Once a set of these 3D volumes is created they can be imported into the cow in one go by having them all in the same folder.

To systematize the conformational flexibility given by the set of 3D Volumes we will use the Principal Component Analysis (PCA) logic. This procedure is heavily depended on the number of eigenvolumes you want to create and the boxsize of your 3D classes. For a standard PC we recomment to calculate less than 20 Eigenvolumes in volumes no bigger than 100 x 100 x 100 voxels. To achieve this the volumes can be downsampled within the cow.

The result of the PCA is a stack of eigenimages, including the average structure and the applied mask as prelast and last item.

To visualize the trajectories we have to calculate the linear factors for each 3D