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Nearly 50 years in the making: defining the catalytic mechanism of the multifunctional enzyme, pyruvate carboxylase
Author(s) -
Menefee Ann L.,
Zeczycki Tonya N.
Publication year - 2014
Publication title -
the febs journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.981
H-Index - 204
eISSN - 1742-4658
pISSN - 1742-464X
DOI - 10.1111/febs.12713
Subject(s) - chemistry , pyruvate carboxylase , mechanism (biology) , context (archaeology) , enzyme kinetics , substrate (aquarium) , steady state (chemistry) , carboxylation , kinetics , enzyme , catalysis , stereochemistry , computational chemistry , active site , biochemistry , physics , biology , paleontology , ecology , quantum mechanics
Numerous steady‐state kinetic studies have examined the complex catalytic reaction mechanism of the multifunctional enzyme, pyruvate carboxylase ( PC ). Through initial velocity, product inhibition, isotopic exchange and alternate substrate experiments, early investigators established that PC catalyzes the Mg ATP ‐dependent carboxylation of pyruvate by HCO 3 – through a nonclassical sequential Bi Bi Uni Uni reaction mechanism. This review surveys previous steady‐state kinetic investigations of PC and evaluates the proposed hypotheses concerning the overall catalytic mechanism, nonlinear kinetics and active site coupling in the context of recent structural and mutagenic analyses of this multifunctional enzyme. The determination several PC holoenzyme structures have aided in corroborating the proposed molecular mechanisms by which catalysis occurs and established the inextricable link between the dynamic protein motions and complex kinetic mechanisms associated with PC activity. Unexpectedly, the conclusions drawn from these early steady‐state kinetic investigations have consistently proven to be in fundamental agreement with our current understanding of PC catalysis, which is a testament to the overarching sophistication of the methods pioneered by Michaelis and Menten and further developed by Northrop, Cleland and others.