Use of isotope effects to characterize intermediates in mechanism-based inactivation of dopamine beta-monooxygenase by beta-chlorophenethylamine

Journal Article · Thu Apr 05 04:00:00 EDT 1990 · Journal of Biological Chemistry; (USA)

OSTI ID:6871660

Bossard, M J; Klinman, J P ^[1]

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1. Univ. of California, Berkeley (USA)

A mechanism for beta-chlorophenethylamine inhibition of dopamine beta-monooxygenase has been postulated in which bound alpha-aminoacetophenone is generated followed by an intramolecular redox reaction to yield a ketone-derived radical cation as the inhibitory species. Based on the assumption that the ketone radical is the inhibitory intermediate, an analogous system was predicted and verified. In the present study, the role of alpha-aminoacetophenone as the proposed intermediate in the inactivation by beta-chlorophenethylamine was examined in greater detail. From the interdependence of tyramine and alpha-aminoacetophenone concentrations, ketone inactivation is concluded to occur at the substrate site as opposed to potential binding at the reductant-binding site. Using beta-(2-1H)- and beta-(2-2H)chlorophenethylamine, the magnitude of the deuterium isotope effect on inactivation under second-order conditions has been found to be identical to that observed under catalytic turnover, D(kappa inact/Ki) = D(kappa cat/Km) = 6-7. By contrast, the isotope effect on inactivation under conditions of substrate and oxygen saturation, D kappa inact = 2, is 3-fold smaller than that seen on catalytic turnover, D kappa cat = 6. This reduced isotope effect for inactivation is attributed to a normal isotope effect on substrate hydroxylation followed by an inverse isotope effect on the partitioning of the enol of alpha-aminoacetophenone between oxidation to a radical cation versus protonation to regenerate ketone. These findings are unusual in that two isotopically sensitive steps are present in the inactivation pathway whereas only one is observable in turnover.

• OSTI ID: 6871660
• Journal Information: Journal of Biological Chemistry; (USA), Journal Name: Journal of Biological Chemistry; (USA) Vol. 265:10; ISSN JBCHA; ISSN 0021-9258
• Country of Publication: United States
• Language: English

Related Subjects

550201* -- Biochemistry-- Tracer Techniques
59 BASIC BIOLOGICAL SCIENCES
AMINES
AROMATICS
AUTONOMIC NERVOUS SYSTEM AGENTS
BIOCHEMICAL REACTION KINETICS
BIOLOGICAL PATHWAYS
CARDIOTONICS
CARDIOVASCULAR AGENTS
CHEMICAL REACTIONS
DEUTERIUM
DOPAMINE
DRUGS
ELEMENTS
ENZYME INHIBITORS
ENZYMES
HYDROGEN ISOTOPES
HYDROXY COMPOUNDS
ISOTOPE APPLICATIONS
ISOTOPE EFFECTS
ISOTOPES
KETONES
KINETICS
LIGHT NUCLEI
MOLECULAR STRUCTURE
NEUROREGULATORS
NONMETALS
NUCLEI
ODD-ODD NUCLEI
ORGANIC COMPOUNDS
OXIDOREDUCTASES
OXYGEN
OXYGENASES
PHENOLS
POLYPHENOLS
RADICALS
REACTION KINETICS
REDOX REACTIONS
STABLE ISOTOPES
SYMPATHOMIMETICS
TRACER TECHNIQUES
TYRAMINE