Oxidized bases in the ribosome's peptidyl transferase center and their effects on translation

Willi, Jessica (23 January 2015). Oxidized bases in the ribosome's peptidyl transferase center and their effects on translation (Unpublished). In: Swiss RNA Workshop 2015. Bern. 23.01.2015.

The ribosome is central to protein biosynthesis and the focus of extensive research. Recent biochemical and structural studies, especially detailed crystal structures and high resolution Cryo-EM in different functional states have broadened our understanding of the ribosome and its mode of action. However, the exact mechanism of peptide bond formation and how the ribosome catalyzes this reaction is not yet understood. Also, consequences of direct oxidative stress to the ribosome and its effects on translation have not been studied. So far, no conventional replacement or even removal of the peptidyl transferase center's bases has been able to affect in vitro translation. Significant contribution to the catalytic activity seems to stem from the ribose-phosphate backbone, specifically 2'OH of A2451. Using the technique of atomic mutagenesis, novel unnatural bases can be introduced to any desired position in the 23S rRNA, surpassing conventional mutagenesis and effectively enabling to alter single atoms in the ribosome. Reconstituting ribosomes in vitro using this approach, we replaced universally conserved PTC bases with synthetic counterparts carrying the most common oxidations 8-oxorA, 5-HOrU and 5-HOrC. To investigate the consequent effects on translation, the chemically engineered ribosomes were studied the in various functional assays. Incorporation of different oxidized bases into the 70S ribosome affected the ribosomes in different ways. Depending on the nucleobase modified, the reconstituted ribosomes exhibited radical deceleration of peptide bond formation, decrease of synthesis efficiency or even an increase of translation rate. These results may further our understanding of the residues involved in the peptide bond formation mechanism, as well as the disease-relevant effects of oxydative stress on the translation machinery.

Item Type:

Conference or Workshop Item (Poster)

Division/Institute:

08 Faculty of Science > Departement of Chemistry and Biochemistry

Graduate School:

Graduate School for Cellular and Biomedical Sciences (GCB)

UniBE Contributor:

Willi, Jessica

Subjects:

500 Science > 570 Life sciences; biology
500 Science > 540 Chemistry

Language:

English

Submitter:

Christina Schüpbach

Date Deposited:

28 Jan 2016 16:17

Last Modified:

28 Jan 2016 16:17

URI:

https://boris.unibe.ch/id/eprint/74841

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