A Ribosome Interaction Surface Sensitive to mRNA GCN Periodicity.

Kristen Scopino,Daniel Krizanc,Elliot Williams,Kelly M Thayer,William A Barr,Abdelrahman Elsayed,Michael P Weir

doi:10.3390/biom10060849

Abstract

A longstanding challenge is to understand how ribosomes parse mRNA open reading frames (ORFs). Significantly, GCN codons are over-represented in the initial codons of ORFs of prokaryote and eukaryote mRNAs. We describe a ribosome rRNA-protein surface that interacts with an mRNA GCN codon when next in line for the ribosome A-site. The interaction surface is comprised of the edges of two stacked rRNA bases: the Watson–Crick edge of 16S/18S rRNA C1054 and the adjacent Hoogsteen edge of A1196 (Escherichia coli 16S rRNA numbering). Also part of the interaction surface, the planar guanidinium group of a conserved Arginine (R146 of yeast ribosomal protein Rps3) is stacked adjacent to A1196. On its other side, the interaction surface is anchored to the ribosome A-site through base stacking of C1054 with the wobble anticodon base of the A-site tRNA. Using molecular dynamics simulations of a 495-residue subsystem of translocating ribosomes, we observed base pairing of C1054 to nucleotide G at position 1 of the next-in-line codon, consistent with previous cryo-EM observations, and hydrogen bonding of A1196 and R146 to C at position 2. Hydrogen bonding to both of these codon positions is significantly weakened when C at position 2 is changed to G, A or U. These sequence-sensitive mRNA-ribosome interactions at the C1054-A1196-R146 (CAR) surface potentially contribute to the GCN-mediated regulation of protein translation.

Highlights

Ratcheting movements and conformational changes of the ribosome large and small subunits are hypothesized to facilitate the threading of mRNA through the ribosome so that its codons can be translated into the protein product
Using molecular dynamics (MD) simulations and molecular modelling, we studied the neighborhood of the mRNA entrance tunnel that the mRNA encounters just before reaching the decoding center A-site
We found that the C nucleotide at the second position of the +1 codon has a specific Watson–Crick/Hoogsteen [14] base pairing with 18S rRNA A1427 (Escherichia coli 16S rRNA A1196, in helix 34) which base stacks with C1054 and is highly conserved as A in 16S and 18S rRNA

Summary

Introduction

Ratcheting movements and conformational changes of the ribosome large and small subunits are hypothesized to facilitate the threading of mRNA through the ribosome so that its codons can be translated into the protein product. In addition to the ratcheting rotation of the small subunit relative to the large subunit, swiveling of the head region relative to the main body/platform of the small subunit is thought to facilitate movement of the tRNA and mRNA from the Aminoacyl-tRNA (A) site (decoding center) to the Peptidyl-tRNA (P) site of the ribosome [1,2]. CCrryyoo--EEMM ssttuuddiieess hhaavvee ssuuggggeesstteedd tthhaatt tthhee yyeeaasstt rriibboossoommee hhaass sseevveerraall iinntteerrmmeeddiiaattee ssttaaggeess ooff ttrraannssllooccaattiioonn rraattcchheettiinngg. CClluusstteerriinngg aannaallyyssiiss ooff vveerryy llaarrggee nnuummbbeerrss ooff ccrryyoo--EEMM iimmaaggeess rreevveeaalleedd fifivvee ddoommiinnaanntt iinntteerrmmeeddiiaattee ssttaaggeess ((ssttaaggeess II tthhrroouugghh VV;; [[55]])). Tghreotraatticohneatinngglerodteactrioeansaesngplreodgreecsrseiavseelys pfrroomgrsetsasgiveeslIytforoVm(1s0t◦a,g5e◦s, 5I ◦t,o1V◦, (01.05°◦,)5p°a, i5r°e,d1w°, i0t.h5°c)hpaanigreesdiwn itthhechheaandgesws iinvetlhaenhgelaed(1s2w◦,iv1e7l◦a, n17g◦le, 1(41◦2,°,11◦)7°[5, ]1.7°, 14°, 1°) [5]

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Conclusion