Naslov
Amber suppression in bacterium Escherichia coli using archaeal ortologous seryl-tRNA

Autori
Lesjak, Sonja ; Boko, Draško ; Rokov-Plavec, Jasmina ; Weygand-Đurašević, Ivana

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

Izvornik
Knjiga sažetaka Kongresa Hrvatskog društva za biokemiju i molekularnu biologiju 2004 / Dumić, Jerka - Zagreb : Hrvatsko društvo za biokemiju i molekularnu biologiju (HDBMB), 2004, 81-81

Skup
Kongres Hrvatskog društva za biokemiju i molekularnu biologiju 2004

Mjesto i datum
HOC Bjelolasica, Hrvatska, 30.09.2004. - 02.10.2004

Vrsta sudjelovanja
Poster

Vrsta recenzije
Domaća recenzija

Ključne riječi
tRNASer ; seryl-tRNA synthetase ; toxicity

Sažetak
Fidelity of translation depends upon aminoacyl- tRNA synthetases, a family of enzymes that recognize cognate tRNAs and charge them with the proper amino acid and are, therefore, essential for any organism. Methanogenic archaeal seryl-tRNA synthetases (SeRSs) have highly unusuall sequences with insertions and deletions, most surprisingly a deletion in motif 2 of the active site. In vitro, methanogenic archaeal SerRSs have shown relative flexibility in recognition of tRNAsSer from all three domains of life, while, on the other hand, Esherichia coli SerRS discriminates strongly against non-homologous tRNAsSer. Based on in vitro assays, we tried to demonstrate the complementation of function of thermosensitive E. coli SerRS (strain KL229) by introducing genes coding for SerRS from Methanocaldococcus jannaschii, Methanopyrus kandleri and Methanococcus maripaludis. Interesting feature of the experiment was the faster growth of colonies transformed with empty vector compared to colonies with vectors carrying genes for archaeal SerRSs on permissive temperature, implying some kind of toxic effect. Because archaeal SerRSs would not complement the function of E. coli SerRS, we wanted to detect the interaction of archaeal SerRSs with cognate tRNAs. Archaeal amber suppressor tRNA genes alongside archaeal SerRS genes were introduced into E. coli strains (JR104 ; XAC-A24) carrying amber mutations. In vivo, suppression was not detected and, again, faster-growing colonies were the ones transformed with empty vectors. This observation was a strong indication of toxic effect. b-galactosidase activity assay performed on strain XAC-A24 has shown that the level of supression does not exceed 10 %. It was suspected that archaeal SerRSs charge noncognate tRNAs with serine in heterologous environment. Accordingly, bacterial tRNATyr was significantly serylated by M. maripaludis SerRS, indicating that archaeal SerRSs in E. coli, in addition to cognate tRNA, might serylate endogenous tRNATyr, thus affecting the fidelity of translation and resulting in synthesis.

Izvorni jezik
Engleski

Znanstvena područja
Kemija, Biologija

POVEZANOST RADA