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Use of isotope effects to determine enzyme mechanisms
Author(s) -
Cleland W. W.
Publication year - 2007
Publication title -
journal of labelled compounds and radiopharmaceuticals
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.432
H-Index - 47
eISSN - 1099-1344
pISSN - 0362-4803
DOI - 10.1002/jlcr.1390
Subject(s) - chemistry , kinetic isotope effect , aspartate carbamoyltransferase , enzyme , stereochemistry , biochemistry , allosteric regulation , quantum mechanics , physics , deuterium
Isotope effects are a powerful tool for determining the mechanisms of enzymatic reactions. Methods for measurement of isotope effects are reviewed and the equations that describe observed isotope effects and permit determination of intrinsic isotope effects on the isotope‐sensitive step are presented. Aspartate transcarbamoylase is used as an example for how the kinetic mechanism can be determined by observing the size of an isotope effect as a function of the concentration of another substrate. Cytidine, adenosine and AMP deaminases are used to illustrate determining the relative rates of steps in the mechanism. Determination of chemical mechanism is illustrated by data for malic enzyme, OMP decarboxylase, aspartate transcarbamoylase, L ‐ribulose‐5‐P 4‐epimerase, oxalate decarboxylase, tryptophan 2‐monooxygenase and the K58A mutant of aspartate aminotransferase when rescued by ammonia. Determination of transition state structure is illustrated by data for formate dehydrogenase, prephenate dehydrogenase, chorismate mutase, as well as for enzymes that catalyze phosphoryl and acyl transfer. Copyright © 2007 John Wiley & Sons, Ltd.