Directional transition from initiation to elongation in bacterial translation

Nucleic Acids Research
Akanksha GoyalM V Rodnina

Abstract

The transition of the 30S initiation complex (IC) to the translating 70S ribosome after 50S subunit joining provides an important checkpoint for mRNA selection during translation in bacteria. Here, we study the timing and control of reactions that occur during 70S IC formation by rapid kinetic techniques, using a toolbox of fluorescence-labeled translation components. We present a kinetic model based on global fitting of time courses obtained with eight different reporters at increasing concentrations of 50S subunits. IF1 and IF3 together affect the kinetics of subunit joining, but do not alter the elemental rates of subsequent steps of 70S IC maturation. After 50S subunit joining, IF2-dependent reactions take place independent of the presence of IF1 or IF3. GTP hydrolysis triggers the efficient dissociation of fMet-tRNA(fMet) from IF2 and promotes the dissociation of IF2 and IF1 from the 70S IC, but does not affect IF3. The presence of non-hydrolyzable GTP analogs shifts the equilibrium towards a stable 70S-mRNA-IF1-IF2-fMet-tRNA(fMet) complex. Our kinetic analysis reveals the molecular choreography of the late stages in translation initiation.

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Citations

Mar 15, 2016·Biopolymers·Cristina Maracci, Marina V Rodnina
May 21, 2016·Protein Science : a Publication of the Protein Society·Marina V Rodnina
Nov 3, 2016·Proceedings of the National Academy of Sciences of the United States of America·Miri KrupkinAda Yonath
Apr 18, 2018·Cold Spring Harbor Perspectives in Biology·Marina V Rodnina
Jun 14, 2017·Molecular and Cellular Biology·Leszek WawiórkaMarek Tchórzewski
Jan 30, 2020·PLoS Biology·Daria S VinogradovaPohl Milón
Sep 20, 2019·Nature·Jinfan WangJoseph D Puglisi
Apr 11, 2018·Proceedings of the National Academy of Sciences of the United States of America·Irena AndreevaMarina V Rodnina
May 18, 2018·Nature Chemical Biology·Ivan L VolkovMagnus Johansson
Apr 25, 2018·Proceedings of the National Academy of Sciences of the United States of America·Xueliang GeSuparna Sanyal
Jul 20, 2019·Proceedings of the National Academy of Sciences of the United States of America·Cédric RomillyE Gerhart H Wagner
Nov 18, 2020·The Journal of Biological Chemistry·Sunil Shetty, Umesh Varshney
Mar 2, 2021·Frontiers in Microbiology·Elena M MaksimovaAndrey L Konevega
Jan 19, 2021·The Journal of Biological Chemistry·Sunil Shetty, Umesh Varshney
May 29, 2021·The Journal of Biological Chemistry·Eva AbsmeierMarkus C Wahl
Jun 24, 2021·Nucleic Acids Research·Jose A NakamotoPohl Milón
Jul 16, 2021·Frontiers in Microbiology·Victor BarrenecheaPohl Milón
Nov 9, 2018·Biochemistry·Ivan L Volkov, Magnus Johansson
Aug 4, 2021·Annual Review of Microbiology·Gert BangeWieland Steinchen

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Methods Mentioned

BETA
FRET
light scattering
GTPase
nucleotide exchange

Software Mentioned

Prism Graphpad
KinTek Explorer

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