Talk abstracts

Talk on Monday 04:45-05:00pm submitted by Sean McClory

Insights into the accurate translation of the genetic code by the ribosome

Sean McClory (OSBP), Aishwarya Devaraj (OSBP), Josh Leisring (Dept. of Microbiology), Daoming Qin (OSBP), Kurt Fredrick (OSBP, Dept. of Microbiology)

Abstract:
In order to synthesize functional proteins from mRNA, the ribosome must correctly pair each 3 base mRNA codon with the corresponding 3 base anticodon of the cognate (correct) aminoacyl(aa)-tRNA. While this process largely depends upon codon-anticodon base-pairing, the ribosome’s job is complicated by the fact that there exists a large cellular pool of near-cognate aa-tRNAs capable of forming partial matches with the codon being translated, and these vastly outnumber cognate aa-tRNAs. The ribosome maintains high accuracy during translation by breaking the aa-tRNA selection process into two phases, initial selection and proofreading, increasing the accuracy by providing two opportunities for near-cognate aa-tRNA to be rejected. These two phases are separated by the irreversible hydrolysis of GTP by elongation factor (EF)-Tu, which delivers the aa-tRNA to the ribosome. Many potent antibiotics target aa-tRNA selection and increase translation errors.
To investigate the mechanism of aa-tRNA selection we used a genetic screen to isolate 34 mutations in the 16S ribosomal (r)RNA that increase translation errors. These mutations cluster around the periphery of the shoulder domain of the small 30S ribosomal subunit, suggesting that a conformational change in this domain is critical for aa-tRNA selection. In vitro, these mutations increase the rate of EF-Tu GTP hydrolysis, similar to error inducing antibiotics. The locations of many mutations are distinct from the binding sites of these antibiotics and may suggest additional areas for drug targets. Some mutations also cause defects in ribosomal proofreading, but others do not, suggesting that the initial selection and proofreading phases of tRNA selection may be governed by distinct mechanisms. These findings challenge several conventional beliefs about the mechanism of this ubiquitous process. Additional characterization of these mutations may provide new insights into how the ribosome maintains the accuracy of translation.

Keywords: translation, ribosome, tRNA