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Thermodynamic characterization of five key kinetic parameters that define neuronal nitric oxide synthase catalysis
Author(s) -
Haque Mohammad Mahfuzul,
Tejero Jesús,
Bayachou Mekki,
Wang ZhiQiang,
Fadlalla Mohammed,
Stuehr Dennis J.
Publication year - 2013
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.12404
Subject(s) - chemistry , catalysis , heme , nitric oxide synthase , kinetic energy , ferric , enzyme , kinetics , hemeprotein , dissociation (chemistry) , atp synthase , ferrous , stereochemistry , inorganic chemistry , biochemistry , organic chemistry , physics , quantum mechanics
NO synthase ( NOS ) enzymes convert l ‐arginine to NO in two sequential reactions whose rates ( k cat1 and k cat2 ) are both limited by the rate of ferric heme reduction ( k r ). An enzyme ferric heme– NO complex forms as an immediate product complex and then undergoes either dissociation (at a rate that we denote as k d ) to release NO in a productive manner, or reduction ( k r ) to form a ferrous heme– NO complex that must react with O 2 (at a rate that we denote as k ox ) in a NO dioxygenase reaction that regenerates the ferric enzyme. The interplay of these five kinetic parameters ( k cat1 , k cat2 , k r , k d and k ox ) determines NOS specific activity, O 2 concentration response, and pulsatile versus steady‐state NO generation. In the present study, we utilized stopped‐flow spectroscopy and single catalytic turnover methods to characterize the individual temperature dependencies of the five kinetic parameters of rat neuronal NOS . We then incorporated the measured kinetic values into computer simulations of the neuronal NOS reaction using a global kinetic model to comprehensively model its temperature‐dependent catalytic behaviours. The results obtained provide new mechanistic insights and also reveal that the different temperature dependencies of the five kinetic parameters significantly alter neuronal NOS catalytic behaviours and NO release efficiency as a function of temperature.