CRYSTALLOGRAPHIC STUDIES OF SIGMA70 SUBUNIT OF E COLI RNA POLYMERASE

University Of Calif-Los Alamos Nat Lab, Los Alamos NM

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Abstract

The SIR provided L-Telluromethionine; 1.5g Our long term goal is to understand the mechanism of transcription and its regulation. Determining the structure of RNA polymerase (RNAP), the enzyme responsible for RNA synthesis, is an essential step towards this goal. This is best accomplished with the highly characterized Escherichia coli RNAP. This is especially true because of the remarkable degree of conservation of RNAP structure and mechanism from bacteria to man. One part of our strategy is to determine high-resolution X-ray structures of RNAP components. We would like to use the same methods we use to prepare selenomthionine protein to prepare telluromethionine substituted protein. This could help us in two ways: 1) If the result of Boles et al. (Nature Structural Biology 1:283-284, 1994) is a general one in that only buried methionine residues are substituted in the final product, then this would likely reduce the number of sites in our protein, making solving for the sites possible 2) Even if the result of Boles et al. is not general and we obtain protein substituted at 10 sites with telluromethionine, the larger isomorphous signal from tellurium (because of the larger number of protons) would likely allow us to solve for the sites.