TAILIEUCHUNG - Báo cáo khóa học: Damped oscillatory hysteretic behaviour of butyrylcholinesterase with benzoylcholine as substrate

Steady-state kinetics for the hydrolysis of benzoylcholine (BzCh) and benzoylthiocholine (BzSCh) by wild-type humanbutyrylcholinesterase (BuChE) andby theperipheral anionic site mutant D70G were /Kmfor the hydrolysis of BzSChwas 17-foldand32-fold lower than that for hydrolysis of BzChbywild-type andD70G, respectively. The rate-limiting step for hydrolysis of BzCh was deacyla-tion, whereas acylation was rate-limiting for hydrolysis of BzSCh. | Eur. J. Biochem. 271 220-234 2004 FEBS 2003 doi Damped oscillatory hysteretic behaviour of butyrylcholinesterase with benzoylcholine as substrate Patrick Masson1 Boris N. Goldstein2 Jean-Claude Debouzy3 Marie-Therese Froment1 Oksana Lockridge4 and Lawrence M. Schopfer4 1 Centre de Recherches du Service de Sante des Armees CRSSA Departement de Toxicologie Unite d Enzymologie La Tronche France 2Institute of Theoretical and Experimental Biophysics Russian Academy of Sciences Moscow Russia 3CRSSA Unite de Biophysique La Tronche France 4University of Nebraska Medical Center Eppley Institute Omaha NE USA Steady-state kinetics for the hydrolysis of benzoylcholine BzCh and benzoylthiocholine BzSCh by wild-type human butyrylcholinesterase BuChE and by the peripheral anionic site mutant D70G were compared. kcat Km for the hydrolysis of BzSCh was 17-fold and 32-fold lower than that for hydrolysis of BzCh by wild-type and D70G respectively. The rate-limiting step for hydrolysis of BzCh was deacylation whereas acylation was rate-limiting for hydrolysis of BzSCh. Wild-type enzyme and the D70G mutant were found to reach steady-state velocity slowly with BzCh as the substrate. At pH 6 the approach to steady-state for both enzymes consisted of a mono-exponential acceleration upon which a set of damped oscillations was superimposed. From pH 7 to the approach to steady-state consisted of a simple exponential acceleration. The damped oscillations were analyzed by both a numerical approximation and simulation based on a theoretical model. BuChE-catalyzed hydrolysis of the thiocholine analogue of BzCh showed neither lags nor oscillations under the same conditions. The frequency and amplitude of the damped oscillations decreased as the BzCh concentration increased. The apparent induction time for the exponential portion of the lag was calculated from the envelope of the damped oscillations or from the smooth lag. Wild-type BuChE showed a hyperboiic .

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