BIOL 98 |
| The tryptophanyl-tRNA synthetase from the thermophilic archaebacteria Methanococcus jannaschii has been cloned and characterized in vitro. The enzyme has been successfully over expressed in Escherichia coli and purified to homogeneity. The enzyme is a dimeric protein composed of 43kD monomers that have an apparent molecular weight of 61kD via polyacrylamide gel electrophoresis. The interaction of the tRNA synthetase and its cognate tRNA has been investigated. The Methanococcus jannaschii tryptophanyl tRNA contains an unusually large dihydrouridine loop that consists of thirteen nucleotides. The important nucleotide residues for aminoacylation by the tryptophan tRNA synthetase have been probed by site directed mutagenesis of the tRNA. In agreement with the tryptophan tRNA synthetases previously characterized from both bacteria and eukaryotes, the discriminator base A73 was found to be important for aminoacylation as well as the anticodon nucleotides C34, C35, and A36. Nucleotides found in the acceptor stem and the unusual dihydrouridine loop and their effects on the rate of aminoacylation will also be discussed. The cross-species aminoacylation of the Methanococcus jannaschii tryptophanyl tRNA by the tryptophanyl tRNA synthetases from Saccharomyces cerevisiae and Escherichia coli was also investigated to provide insight into the tRNA identity elements important for maintaining species specific aminoacylation by the tRNA synthetases from the three representative orders of life. |
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Chemistry and Metabolism
4:30 PM-6:30 PM, Tuesday, 12 September 2006 Moscone Center -- Hall D, Poster
Division of Biological Chemistry |