PMID: 764868Feb 24, 1976

Mechanism of aminoacylation of tRNA. Proof of the aminoacyl adenylate pathway for the isoleucyl- and tyrosyl-tRNA synthetases from Escherichia coli K12

Biochemistry
A R Fersht, M M Kaethner

Abstract

The following observations show that the formation of isoleucyl-tRNA catalyzed by the isoleucyl-tRNA synthetase from Escherichia coli K12 involves the initial rapid formation of an isoleucyl adenylate complex followed by the slow, rate-determining, transfer of the isoleucyl moiety to tRNA. (1) The rate constant for the transfer of [14C]Ile from the E-[14C]Ile approximately AMP complex to tRNA is the same as the turnover number for the steady-state isoleucylation of tRNA at pH 7.78 (1.5 s-1) and pH 5.87 (0.34 s-1). (2) On mixing a solution of isoleucyl-tRNA synthetase and tRNA with [14C]Ile and ATP the steady-state rate of isoleucylation is attained in the first turnover of the enzyme, with little or no "burst" or charging that would indicate a slow step after the transfer step. (3) The pyrophosphate exchange reaction in the presence of tRNA is 40 times faster than the overall rate of isoleucylation of tRNA. (4) Similarly, rapid quenching experiments indicate that isoleucyl adenylate is formed prior to the transfer step. The possibility that isoleucyl adenylate formation is a parallel reaction caused by a second active site on the enzyme is ruled out both by the stoichiometry in this rapid quenching experiment and also the overa...Continue Reading

References

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May 10, 1980·Nucleic Acids Research·C L Harris, F Marashi
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Jan 29, 2002·Archives of Biochemistry and Biophysics·Matthew G GuentherJames R Burke
Jul 8, 2011·Organic & Biomolecular Chemistry·Raj S DhimanRonald Kluger
Mar 15, 1977·FEBS Letters·R S Taylor, A G Weeds
Feb 5, 2008·Methods : a Companion to Methods in Enzymology·Christopher S FrancklynYa-Ming Hou
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Sep 11, 2012·The Journal of Physical Chemistry. B·Mantu Santra, Biman Bagchi

Citations

Nov 1, 1976·FEBS Letters·R JakesJ Kendrick-Jones
Jul 16, 1973·European Journal of Biochemistry·A DurekovićM R Kula
Sep 1, 1970·European Journal of Biochemistry·F Berthelot, M Yaniv

Related Concepts

Amino Acid Activation, Translational
Alkalescens-Dispar Group
Transfer RNA
Alloisoleucine
Amino Acyl-tRNA Synthetases
Phosphaden
IARS
Tyrosine
Isoleucine-tRNA Ligase
RNA, Bacterial

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