RNA Polymerase Flexing and Animations

A flexible alignment tool was recently added to our 'Situs' docking package that is based on 3D 'motion capture' technology used in the entertainment industry and in biomechanics. We have used this tool in collaboration with Seth Darst (Rockefeller University), whose laboratory determined the structure of Escherichia coli core RNA polymerase (RNAP) by cryo-electron microscopy and image processing of helical crystals to a resolution of 15Å. Because of the high sequence conservation between the core RNAP subunits, we were able to interpret the E. coli structure in relation to the high-resolution X-ray structure of Thermus aquaticus core RNAP. A very large conformational change of the T. aquaticus RNAP X-ray structure, corresponding to opening of the main DNA/RNA channel by nearly 25Å, was required to fit the E. coli map (see Figure). This finding reveals, at least partially, the range of conformational flexibility of the RNAP, which is likely to have functional implications for the initiation of transcription, where the DNA template must be loaded into the channel.

Figure: Direct space flexible fitting with skeletons. This overview illustrates the skeleton-based modeling steps that were employed in the recent flexible refinement of the T. aquaticus crystal structure (upper right) of RNA polymerase against EM data (upper left) from the laboratory of Seth Darst. The flexed model (bottom) is compared to the EM data (bottom right). By freezing inessential degrees of freedom, skeletons of connected landmarks significantly reduce the effect of noise and thereby improve the stereochemical quality of the fitted structures relative to unconstrained alignments.


Quicktime Movies: [ view 1 | view 2 | view 3 ]

These animations created by Seth Darst demonstrate the conformational change induced by the transition from the X-ray structure to the EM envelope. The color codes for the actual displacement, where warm colors move more than cold colors.


Main Page