Naslov
Interplay between isoleucyl-tRNA synthetase and tRNAIle for optimized amino acid recognition in translation

Autori
Cvetešić, Nevena ; Biluš, Mirna ; Gruić-Sovulj, Ita

Vrsta, podvrsta i kategorija rada
Sažeci sa skupova, sažetak, znanstveni

Izvornik
10th International Symposium on Aminoacyl-tRNA Synthetases / - , 2015, 21-21

Skup
10th International Symposium on Aminoacyl-tRNA Synthetases

Mjesto i datum
Barcelona, Španjolska, 18.10.2015. - 22.10.2015

Vrsta sudjelovanja
Predavanje

Vrsta recenzije
Nije recenziran

Ključne riječi
aminoacyl-tRNA synthetase; isoleucyl-tRNA synthetase; proofreading; ribonucleoprotein

Sažetak
Escherichia coli isoleucyl-tRNA synthetase (IleRS) is distinguished by its use of robust tRNA-dependent pre-transfer editing to eliminate the misactivated valyl-AMP intermediate. We have recently shown that a synthetic site residue tyrosine 59 participates in amino acid activation and transfer step, and acts as a key determinant for tRNA-dependent pre-transfer editing. This clearly demonstrates that the tRNA-dependent synthetic and editing pathways are interwoven within the IleRS synthetic site. Further, IleRS may use tRNA to modulate amino acid activation step [2]. To provide insight into the mechanism by which tRNAIle coordinates synthesis vs hydrolysis of the aminoacyl-adenylate within the synthetic site, detailed kinetic analyses of the synthetic and editing pathways using several tRNAIle variants were performed [3]. These variants encompass the native tRNAIle, and tRNAs lacking the A76 2’OH or the A76 3’OH terminal group. Our results demonstrate that the presence of tRNAIle affects both the synthetic and the hydrolytic reaction. The IleRS:tRNAIle complex exhibits a 10-fold drop in amino acid affinity relative to the free enzyme, and a 10-fold faster aminoacyl- adenylate hydrolysis regardless of the tRNAIle variant present in the complex. By using tRNA lacking the A76 2’OH nucleophile, we uncoupled aminoacylation and tRNA-dependent pre-transfer editing thus demonstrating for the first time that the A76 hydroxyl groups participate only in post-transfer editing. Intriguingly, the discrimination against valine was not altered by the presence of tRNA as both the affinity and pre-transfer editing were equally modulated regardless of the amino acid identity. It appears that the amino acid specificity is ensured by the protein component, while the tRNA adjusts the catalytic site to match the enzyme’s affinity with the cellular amino acid concentration. Without this modulation, aminoacylation and editing would occur almost equally often, as IleRS would operate near saturation with both the cognate isoleucine and the non-cognate valine under physiological conditions. We propose that IleRS acts as a ribonucleoprotein to minimise the cost of energetically expensive editing through optimised amino acid recognition in the cellular context.

Izvorni jezik
Engleski

Znanstvena područja
Kemija, Biologija

POVEZANOST RADA