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'''Liu, H , Yap, LP , Stehlin, C | '''Liu, H, Yap, LP, Stehlin, C and Musier-Forsyth, K''' (1995) Molecular recognition of tRNA(Pro) by Escherichia coli proline-tRNA synthetase. ''Nucleic Acids Symp. Ser.'' '''''':176-8 | ||
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Latest revision as of 04:55, 15 September 2013
| Citation |
Liu, H, Yap, LP, Stehlin, C and Musier-Forsyth, K' (1995) Molecular recognition of tRNA(Pro) by Escherichia coli proline-tRNA synthetase. Nucleic Acids Symp. Ser. ':176-8 |
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| Abstract |
We have investigated the molecular recognition of tRNA(Pro) by Escherichia coli proline tRNA synthetase (ProRS) in vitro using semi-synthetic tRNAs and site-directed mutagenesis of full-length tRNA transcripts. These studies have led to an improved understanding of how this class II synthetase interacts with its tRNA substrate. The ability to efficiently aminoacylate a tRNA(Pro) molecule assembled by annealing together a shorter, chemically synthesized oligonucleotide and a 3/4 tRNA prepared enzymatically, has facilitated the identification of RNA structural features that are critical for aminoacylation by ProRS. This approach has been successful using either a 3'-3/4 tRNA annealed to a 5'-oligonucleotide or a 5'-3/4 tRNA annealed to a 3'-oligonucleotide. These studies show that ProRS appears to be particularly sensitive to mutations that result in structural changes in the core region of tRNA(Pro). Moreover, the so-called "variable pocket" nucleotides appear to be dispensable for aminoacylation. We have also identified a specific 2'-hydroxyl-base interaction between the ribose of U8 and the 2-amino group of G46 that makes a thermodynamically significant contribution to tRNA(Pro) aminoacylation by E. coli ProRS. |
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| Keywords |
Amino Acyl-tRNA Synthetases/metabolism; Base Sequence; Binding Sites; Escherichia coli/genetics; Escherichia coli/metabolism; Hydroxylation; Molecular Sequence Data; Molecular Structure; Mutagenesis, Site-Directed; Nucleic Acid Conformation; RNA, Transfer, Pro/chemistry; RNA, Transfer, Pro/genetics; RNA, Transfer, Pro/metabolism; Thermodynamics |
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