Talk abstracts

Talk on Friday 01:10-01:25pm submitted by Aishwarya Devaraj

A role for the 30S subunit E site in maintenance of the translational reading frame

Aishwarya Devaraj (Ohio State Biochemistry Program, OSU), Shinichiro Shoji (Microbiology, OSU), Eric D Holbrook (Microbiology, OSU), Kurt Fredrick (Microbiology and Ohio State Biochemistry Program, OSU)

Abstract:
The exit (E) site has been implicated in several ribosomal activities, including translocation, decoding, and maintenance of the translational reading frame. Here, we target the 30S subunit E site by introducing a deletion in rpsG that truncates the beta-hairpin of ribosomal protein S7. This mutation (S7DeltaR77-Y84) increases both -1 and +1 frameshifting but does not increase miscoding, providing evidence that the 30S E site plays a specific role in frame maintenance. Mutation S7DeltaR77-Y84 also stimulates +1 programmed frameshifting during prfB'-lacZ translation in many synthetic contexts. However, no effect is seen when the E codon of the frameshift site corresponds to those found in nature, suggesting that E-tRNA release does not normally limit the rate of prfB frameshifting. Ribosomes containing S7DeltaR77-Y84 exhibit an elevated rate of spontaneous reverse translocation and an increased K (1/2) for E-tRNA. These effects are of similar magnitude, suggesting that both result from destabilization of E-tRNA. Finally, this mutation of the 30S E site does not inhibit EF-G-dependent translocation, consistent with a primary role for the 50S E site in the mechanism

Keywords: Ribosome, E site, Frameshifting