Antioxidant-Mediated Modulation of Protein Reactivity for 3,4-Dihydroxyphenylacetaldehyde, a Toxic Dopamine Metabolite

Abstract

3,4-Dihydroxyphenylacetaldehyde (DOPAL) is an endogenously produced toxic aldehyde. It is a bifunctional electrophile implicated in the loss of dopaminergic cells concomitant with Parkinson’s disease and neurodegeneration. DOPAL is known to react with proteins and amino acids such as N-acetyl Lys; oxidation of the catechol moiety to the quinone of DOPAL increases this reactivity. Here we demonstrate the ability of the antioxidants N-acetylcysteine, glutathione, and ascorbic acid to mitigate the reactivity of DOPAL with proteins and amino acids in a dose-dependent fashion. Conversely, Trolox did not lessen the observed reactivity with proteins. Interestingly, use of tricine, a buffer and reducing agent, in these systems also decreased the reactivity of DOPAL with amines, yielding tricine-derived free radical species. Modification of amines with aldehydes typically involves Schiff base chemistry; however, the observance of free radicals suggests that an oxidative step is involved in the reaction of DOPAL with lysine. Furthermore, while Schiff base formation is usually optimal at pH 5, the reaction rate of DOPAL with N-acetyl Lys is negligible at pH 5 and is enhanced under basic conditions (e.g. pH 9). Conditions of high pH are also favorable for catechol auto-oxidation, known to occur for DOPAL. The antioxidant-mediated protection demonstrated here suggests that oxidative stress may impart cellular vulnerability to protein modification by DOPAL. Therefore, depleted antioxidants and increased levels of lipid peroxidation products, known to prevent detoxifying metabolism of DOPAL, may present a survival challenge to dopaminergic cells targeted in Parkinson’s disease.

3,4-Dihydroxyphenylacetaldehyde-Induced Protein Modifications and Their Mitigation by N-Acetylcysteine

“…Consistent with previous reports that a variety of antioxidants inhibit DOPAL reactivity with proteins (Rees et al, 2009;Follmer et al, 2015), treatment with NAC was remarkably effective in mitigating all the protein modifications exerted by DOPAL-quinonization, oligomerization, ubiquitination, and aggregation-as well as the inhibitory effect of DOPAL on LAAAD. The finding that NAC prevented DOPALinduced AS oligomerization replicates the report by Anderson et al (2016).…”

“…To explore whether protein modifications exerted by DOPAL depend on its oxidation, we assessed effects of the antioxidant N-acetylcysteine (NAC). A variety of antioxidants interfere with DOPAL-induced oligomerization of AS (Follmer et al, 2015;Anderson et al, 2016). We chose NAC for the present experiments for several reasons.…”

“…NAC decreases endogenous DOPAL levels in PC12 cells (Goldstein et al, 2017a) and prevents DA-evoked formation of intracellular quinoprotein adducts (Banerjee et al, 2014). Finally, NAC is known to inhibit DOPAL-induced oligomerization of AS (Anderson et al, 2016).…”

The “Sick-but-not-Dead” Phenomenon Applied to Catecholamine Deficiency in Neurodegenerative Diseases

“…156,163,164,177 In the few studies where DOPAL and dopamine have been compared directly in terms of oligomerizing AS, DOPAL has been found to be more potent. 156,163,171 We recently compared DOPAL and dopamine in terms of (1) AS oligomerization and quinonization in test tube experiments; (2) enhancing effects of Cu(II) 171,178 and mitigating effects of antioxidation with N-acetylcysteine (NAC) 134,156,179,180 ; and (3) quinonization of intracellular proteins in cultured cells. 134,156 We found that DOPAL is far more potent than dopamine in both oligomerizing and quinonizing AS.…”

Toxic dopamine metabolites, oxidative stress, and antioxidants as contributors to neurodegeneration specifically Parkinson’s disease

“…137 Then work by Anderson et al used EPR to detect free radicals during DOPAL's interaction with N-acetyl lysine, an amino acid residue known to interact with DOPAL. 138 It was expected that the Schiffbase radical that is formed would give a splitting pattern that was comparable to the DOPAL radical, however these splitting patterns were absent. 138 Instead the splitting patterns were consistent with nitroxide free radicals derived from tricine, further supporting the evidence that tricine forms a stable radical.…”