Pyridoxamine:  An Extremely Potent Scavenger of 1,4-Dicarbonyls

Abstract

1,4-Dicarbonyl compounds, which include 2,5-hexanedione and recently discovered endogenous 4-ketoaldehydes (levuglandins, isoketals, and neuroketals), exhibit severe toxicity. The key step in the toxicity of these compounds is their reaction with the lysyl residues of proteins to form pyrrole adducts. To screen for effective scavengers of these toxic compounds, we determined the reaction rates of pyrrole formation for a series of primary amines with a model 4-ketoaldehyde, 4-oxopentanal (OPA). We found pyridoxamine (PM) to react extremely rapidly, with a second-order rate constant at physiological pH being approximately 2300 times faster than that of Nalpha-acetyllysine. The extreme reactivity of PM was unique to 1,4-dicarbonyls, as its reactions with methylglyoxal and 4-hydroxy-2(E)-nonenal were much slower and only slightly faster than with Nalpha-acetyllysine. The phenolic group of PM was found to be essential to its high reactivity, and the rate constant for pyrrole formation with OPA exhibited a maximum at pH 7.5, close to the second pKa of PM. We therefore propose a mechanism involving transfer of the phenolic proton to the carbonyl of the initially formed hemiacetal, which facilitates subsequent nucleophilic attack and ring closure. Only 1,4-dicarbonyls are likely to participate in the proposed mechanism, thereby conferring unique sensitivity of this class of compounds to scavenging by PM.

Pyridoxamine Analogues Scavenge Lipid-Derived γ-Ketoaldehydes and Protect against H2O2-Mediated Cytotoxicity

“…One of the most promising and selective strategies might be to scavenge these γKA with amine containing compounds that are more reactive than lysyl groups. Previous observations suggested that PM was a good candidate for a γKA scavenger because it reacts with γKAs about 2,000 times faster than does lysine (29). Our present study showed that PM significantly inhibited the formation of lysylIsoK adducts when coincubated with excess lysine and oxidized arachidonic acid.…”

“…118-120 °C; MS m/z 239 (M + 1), 222 (M -NH 2 ), 151 (222 -C 5 H 11 ), 136 (151 -CH 3 ). To determine the second order rate constant for pyrrole formation with a model γKA, 4-oxo-pentanal, 1 mM each of 4-oxo-pentanal and PPM, PM, or SA were incubated together and measurements carried out as described in (29) except that the reaction buffer was 50 mM phosphate buffer in 1:1 acetonitrile-water.…”

“…We previously determined that the reaction of pyridoxamine with a model γKA, 4-oxopentanal, is more than 2,000 times faster than that of N α -acetyllysine with 4-oxopentanal (29 …”

“…Therefore, lipophilic analogs of PM may be more useful scavengers of endogenously formed isoketals. Salicylamine, like PM, includes adjacent methylamine and hydroxyl substituents on an aromatic ring, and reacts only slightly slower than PM with γ-ketoaldehydes in PBS (29), but is much more lipophilic because there is no pyridine ring nitrogen. To retain all of the structural determinants of PM, but to increase lipophilicity, we also modified PM by converting the 5'-hydroxyl group to a pentyl ether group to form 5'-Opentylpyridoxamine ( Figure 5).…”

“…Therefore, modification of PM in a way that retained its high reactivity but increases its lipophilicity could enhance its effectiveness as a scavenger. We previously found that salicylamine (SA), which readily dissolves in ethyl acetate, also rapidly reacted with γKAs (29). We interpreted this finding to suggest that an aminomethyl group and an adjacent hydroxyl on an aromatic ring were the critical components for γKA scavenging.…”