Dynamics of RNA modification by a multi-site-specific tRNA methyltransferase. - Laboratoire de Chimie des Processus Biologiques Accéder directement au contenu
Article Dans Une Revue Nucleic Acids Research Année : 2015

Dynamics of RNA modification by a multi-site-specific tRNA methyltransferase.

Résumé

In most organisms, the widely conserved 1-methyl-adenosine58 (m(1)A58) tRNA modification is catalyzed by an S-adenosyl-L-methionine (SAM)-dependent, site-specific enzyme TrmI. In archaea, TrmI also methylates the adjacent adenine 57, m(1)A57 being an obligatory intermediate of 1-methyl-inosine57 formation. To study this multi-site specificity, we used three oligoribonucleotide substrates of Pyrococcus abyssi TrmI (PabTrmI) containing a fluorescent 2-aminopurine (2-AP) at the two target positions and followed the RNA binding kinetics and methylation reactions by stopped-flow and mass spectrometry. PabTrmI did not modify 2-AP but methylated the adjacent target adenine. 2-AP seriously impaired the methylation of A57 but not A58, confirming that PabTrmI methylates efficiently the first adenine of the A57A58A59 sequence. PabTrmI binding provoked a rapid increase of fluorescence, attributed to base unstacking in the environment of 2-AP. Then, a slow decrease was observed only with 2-AP at position 57 and SAM, suggesting that m(1)A58 formation triggers RNA release. A model of the protein-tRNA complex shows both target adenines in proximity of SAM and emphasizes no major tRNA conformational change except base flipping during the reaction. The solvent accessibility of the SAM pocket is not affected by the tRNA, thereby enabling S-adenosyl-L-homocysteine to be replaced by SAM without prior release of monomethylated tRNA.

Domaines

Chimie organique
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Dates et versions

hal-01076766 , version 1 (27-02-2024)

Identifiants

Citer

Djemel Hamdane, Amandine Guelorget, Vincent Guérineau, Béatrice Golinelli-Pimpaneau. Dynamics of RNA modification by a multi-site-specific tRNA methyltransferase.. Nucleic Acids Research, 2015, 42 (18), pp.11697-11706. ⟨10.1093/nar/gku820⟩. ⟨hal-01076766⟩
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