Mechanism for reactivation of N-cyclopropylbenzylamine-inactivated monoamine oxidase by amines

Abstract

The effect of 18 different amines, two mercaptans, and two alcohols on the reactivation of N-cyclopropylbenzylamine- (N-CBA-) inactivated bovine liver monoamine oxidase (MAO) is described. All of the compounds that reactivate the enzyme produce a time-dependent pseudo-first-order return of enzyme activity and exhibit saturation kinetics. There is no direct correlation between the ability of a compound to serve as a substrate for native MAO and its ability to reactivate N-CBA-inactivated MAO. Amines containing an aromatic moiety, in general, are better reactivators than the aliphatic amines. The amine must be primary or secondary in order for reactivation to occur. The distance between the aromatic portion and the amino group is critical to the reactivation properties of the compound. The mercaptans and alcohols do not reactivate N-CBA-inactivated MAO, nor do they interfere with the reactivation reaction by benzylamine. Three mechanisms for the reactivation reaction are considered. One involves initial Schiff base formation with the active site adduct produced by N-CBA inactivation of MAO followed by base-catalyzed beta-elimination to the imine of acrolein. The second mechanism is the same as the first except no prior Schiff base formation is invoked. The third mechanism is an SN2 displacement by the amine of the active site amino acid residue attached to the adduct. Experiments are carried out to exclude the SN2 mechanism. The results of the reactivation experiments favor the Shiff base mechanism.

Revised mechanism for inactivation of mitochondrial monoamine oxidase by N-cyclopropylbenzylamine

“…Since N-CBA is itself a secondary amine, it, too, could release [3H]acrolein. The fcinact at saturation for N-CBA is 461 times greater than the &react of saturation for '-methylbenzylamine, a model secondary amine for N-CBA (Yamasaki & Silverman, 1985); therefore, inactivation would occur much more rapidly than reactivation, and the enzyme would appear to have no activity throughout this process.…”

“…We suspected that the continued generation of [3H] acrolein following inactivation of the enzyme resulted from reactivation and further inactivation of the inactivated enzyme by N-CBA. Primary and secondary amines were shown to produce reactivation of N-CBA-inactivated MAO with concomitant release of acrolein (Yamasaki & Silverman, 1985). Since N-CBA is itself a secondary amine, it, too, could release [3H]acrolein.…”

“…Only 1 equiv of the 3.1 equiv of tritium from N-[1-3H]CBA which was incorporated into MAO during inactivation was released into solution following treatment with the known reactivator benzylamine Yamasaki & Silverman, 1985). Since full reactivation of the enzyme occurred with this released radioactivity, it may be inferred that 1 equiv of inactivator was bound to the active site, whereas the other 2 equiv were peripherally attached.…”

Inactivation of monoamine oxidase A by the monoamine oxidase B inactivators 1-phenylcyclopropylamine, 1-benzylcyclopropylamine, and N-cyclopropyl-.alpha.-methylbenzylamine

“…0006-2960/88/0427-8448S01.50/0 © 1988 American Chemical Society (Silverman & Hoffman, 1980; Vazquez ; Yamasaki & Silverman, 1985), A-(lmethylcyclopropyl)benzylamine (Silverman & Hoffman, 1981; Silverman & Yamasaki, 1984), A-cyclopropyl-a-methylbenzylamine (Silverman, 1984;Vazquez & Silverman, 1985), 1-phenylcyclopropylamine (Silverman & Zieske, 1985a, 1986, and 1-benzylcyclopropylamine (Silverman & Zieske, 1985b), also were determined to be MAO inactivators. All of these inactivation studies suggest that radical intermediates are involved in MAO catalysis and inactivation, and recently a radical intermediate was spin-trapped and observed by ESR spectroscopy (Yelekci & Silverman, unpublished results).…”

A new room temperature molten salt solvent system: organic cation tetrachloroborates

“…A New Room Temperature Molten Salt Solvent System: Organic Cation Tetrachloroborates Received January 12,1987 In the past decade there has been considerable interest in AlCl3-containing molten salts. These melts provide novel media for fundamental studies and are also of interest in high-energy batteries and for catalytic applications.1 Relatively few molten salt systems are liquid at or below room temperature.…”