STRUCTURE DETERMINATION OF THE RNAP ALPHA SUBUNIT

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The long term goal is to understand the regulation mechanism of transcription initiation and elongation of RNA polymerase. The alpha subunit is a critical part of the RNA polymerase from E. coli. It initiates the assembly of the functional enzyme itself. It also serves as a bridge between the enzyme and its external regulators such as many transcription activators, and it contracts DNA as well. Due to the difficulty to obtain high resolution structure of RNA polymerase as a whole, the following approach is the bet way to gain detailed structure information form the complex. That is determining the atomic structure of the alpha subunit by x-ray crystallography and finding the location of alpha subunit in the complex from low resolution electron microscopy structure of RNA polymerase. Low resolution structures have been obtained by Dr. Darst and his co-workers of E. coli RNAP holoenzyme (27 A resolution) and E. coli core enzyme (12 A resolution). Concurrently, experiments are underway using antibody labeling and other methods to locate the alpha subunits within the density of the low-resolution structure. The combined X-ray crystallographic and electron microscopy data will provide more information for thinking about RNAP assembly and function than either method will provide alone.