The structure of the zinc sites of Escherichia coli DNA-dependent RNA polymerase.
X-ray absorption spectroscopy is ideally suited for the investigation of the electronic structure and the local environment (approximately 5 A) of specific atoms in biomolecules. While the edge region provides information about the valence state of the absorbing atom, the chemical identity of neighboring atoms, and the coordination geometry, the extended x-ray absorption fine structure region contains information about the number and average distance of neighboring atoms and their relative disorder. The development of sensitive detection methods has allowed studies using near physiological concentrations (as low as approximately 100 microM). RNA polymerase from Escherichia coli contains two zinc atoms: one tightly bound in the beta' subunit, the subunit that participates in template binding, and the other loosely bound in the beta subunit, the subunit that participates in substrate binding. X-ray absorption studies of these zinc sites in the native protein and of the zinc site in the beta' subunit after removal of the zinc in the beta subunit site by p-(hydroxymercuri)benzenesulfonate (Giedroc, D. P., and Coleman, J. E. (1986) Biochemistry 25, 4969-4978) indicate that both zinc sites have octahedral coordination. The zinc in the beta' subunit site has four sulfur ligands at an average distance of 2.36 +/- 0.02 A and two oxygen (or nitrogen) ligands at an average distance of 2.23 +/- 0.02 A. The beta subunit zinc site has five sulfur ligands at an average distance of 2.38 +/- 0.01 A and one histidine nitrogen ligand at 2.14 +/- 0.02 A. These results are in general agreement with earlier biochemical and spectroscopic studies.
Wu
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Department of Pharmacological Sciences, State University of New York, Stony Brook 11794.
Huang
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Sinclair
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Powers
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D015502:Absorptiometry, Photon / Q000379:methods; D000595:Amino Acid Sequence; D001665:Binding Sites; D012321:DNA-Directed RNA Polymerases / Q000737:chemistry* / Q000378:metabolism*; D004926:Escherichia coli / Q000201:enzymology*; D005583:Fourier Analysis; D007700:Kinetics; D046911:Macromolecular Substances; D008958:Models, Molecular; D008969:Molecular Sequence Data; D011487:Protein Conformation; D015032:Zinc / Q000032:analysis / Q000378:metabolism*
D046911:Macromolecular Substances; D012321:DNA-Directed RNA Polymerases; D015032:Zinc