Naslov
IleRS eliminates norvaline from Escherichia coli proteome via pre- and post-transfer editing pathways

Autori
Biluš, Mirna ; Gruić-Sovulj, Ita

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

Izvornik
Biomolecular complexes and assemblies / Hozić, Amela ; Vuletić, Tomislav - Zagreb, 2014, 56-56

ISBN
978-953-7941-02-4

Skup
12th Greta Pifat Mrzljak International School of Biophysics

Mjesto i datum
Primošten, Hrvatska, 27.09.2014. - 06.10.2014

Vrsta sudjelovanja
Poster

Vrsta recenzije
Nije recenziran

Ključne riječi
isoleucyl-tRNA synthetase; norvaline; proofreading; Escherichia coli

Sažetak
Isoleucyl-tRNA synthetase (IleRS) catalyzes ATP- dependent covalent pairing of cognate isoleucine with cognate tRNAIle, thus providing accurate substrates for ribosomal protein synthesis. IleRS may also efficiently substitute isoleucine with structurally similar valine in the synthetic reaction. To prevent erroneous aminoacylation, IleRS developed extensive hydrolytic proofreading. This enzyme employs both the tRNA-dependent pre- transfer (hydrolysis of Val-AMP intermediate) and the post-transfer (hydrolysis of Val-tRNAIle) editing reactions within the synthetic and editing site, respectively[1]. Norvaline is a natural non- proteinogenic amino acid, structurally similar to isoleucine. It may accumulate in Escherichia coli under hypoxic conditions to milimolar concentrations. We have recently shown that norvaline is the major threat for error-free Leu- tRNALeu synthesis in E. coli[2]. To establish the mechanism of norvaline discrimination by IleRS, we utilized steady state and single-turnover kinetic analyses of the synthetic and editing pathways. We show that norvaline, similarly as valine, is not discriminated well in the synthetic reaction, and is efficiently activated and transferred to tRNAIle. However, the rapid post-transfer editing reaction precludes accumulation of Nva-tRNAIle, acting as the main defense against misincorporation of norvaline in place of isoleucine into cellular proteome. We further demonstrate that tRNA-dependent pre-transfer editing, that efficiently operates against Val- AMP, is also employed by IleRS to hydrolyze Nva- AMP. Our data thus establish that the tRNA- dependent pre-transfer editing activity, an idiosyncratic feature of IleRS proofreading, is characteristic of the IleRS:tRNAIle complex and is independent on the identity of non-cognate amino acid substrate.

Izvorni jezik
Engleski

Znanstvena područja
Kemija, Biologija

POVEZANOST RADA