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PMID: 25246550 Published · ppublish English Journal Article Research Support, Non-U.S. Gov't

Ribosome-induced tuning of GTP hydrolysis by a translational GTPase.

Proceedings of the National Academy of Sciences of the United States of America ·Vol. 111 ·No. 40 ·2014-10-07 ·Pages 14418-23

Maracci C, Peske F, Dannies E, Pohl C, Rodnina MV

Abstract

GTP hydrolysis by elongation factor Tu (EF-Tu), a translational GTPase that delivers aminoacyl-tRNAs to the ribosome, plays a crucial role in decoding and translational fidelity. The basic reaction mechanism and the way the ribosome contributes to catalysis are a matter of debate. Here we use mutational analysis in combination with measurements of rate/pH profiles, kinetic solvent isotope effects, and ion dependence of GTP hydrolysis by EF-Tu off and on the ribosome to dissect the reaction mechanism. Our data suggest that--contrary to current models--the reaction in free EF-Tu follows a pathway that does not involve the critical residue H84 in the switch II region. Binding to the ribosome without a cognate codon in the A site has little effect on the GTPase mechanism. In contrast, upon cognate codon recognition, the ribosome induces a rearrangement of EF-Tu that renders GTP hydrolysis sensitive to mutations of Asp21 and His84 and insensitive to K(+) ions. We suggest that Asp21 and His84 provide a network of interactions that stabilize the positions of the γ-phosphate and the nucleophilic water, respectively, and thus play an indirect catalytic role in the GTPase mechanism on the ribosome.

MeSH Terms
Aspartic Acid/chemistry,genetics,metabolism Binding Sites/genetics Catalysis Catalytic Domain Codon/genetics,metabolism GTP Phosphohydrolases/chemistry,genetics,metabolism Guanosine Triphosphate/chemistry,metabolism Histidine/chemistry,genetics,metabolism Hydrogen-Ion Concentration Hydrolysis Kinetics Models, Molecular Molecular Conformation Molecular Structure Mutation Peptide Elongation Factor Tu/chemistry,genetics,metabolism Protein Binding Protein Biosynthesis RNA, Transfer, Amino Acyl/metabolism Ribosomes/chemistry,metabolism
Chemicals
Codon RNA, Transfer, Amino Acyl Aspartic Acid Histidine Guanosine Triphosphate GTP Phosphohydrolases Peptide Elongation Factor Tu
Authors & Affiliations
5 authors, click to expand affiliations / ORCID
Maracci Cristina
Department of Physical Biochemistry, Max Planck Institute for Biophysical Chemistry, 37077 Göttingen, Germany.
Peske Frank
Department of Physical Biochemistry, Max Planck Institute for Biophysical Chemistry, 37077 Göttingen, Germany.
Dannies Ev
Department of Physical Biochemistry, Max Planck Institute for Biophysical Chemistry, 37077 Göttingen, Germany.
Pohl Corinna
Department of Physical Biochemistry, Max Planck Institute for Biophysical Chemistry, 37077 Göttingen, Germany.
Rodnina Marina V
Department of Physical Biochemistry, Max Planck Institute for Biophysical Chemistry, 37077 Göttingen, Germany rodnina@mpibpc.mpg.de.
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Article Info
Journal
Proceedings of the National Academy of Sciences of the United States of America
Abbr.
Proc Natl Acad Sci U S A
ISSN
1091-6490
DOI
10.1073/pnas.1412676111
Email
rodnina@mpibpc.mpg.de
Published
2014-10-07
Epub
2014-00-22
Pages
14418-23
Language
English
Region
United States
NLM ID
7505876
PMCID
PMC4210003
Subset
IM
External Links
PubMed source DOI: 10.1073/pnas.1412676111 PMC Full Text View journal details
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