Aza-quinone methides (aQMs) are a class of electrophilic intermediates that have thus far been applied to organic synthesis and biological probes and prodrugs. aQMs display a notable chemical similarity to quinone methides (QMs), which are analogous oxygen-containing intermediates known to alkylate numerous biomolecules. However, unlike QMs, the alkylation chemistry of aQMs has received little attention. To generate aQMs, we use the enzyme NfsB which reduces nitroaromatic substrates to their hydroxylamine derivatives forming aQM precursors. These species undergo subsequent 1,6- or 1,4- elimination at the benzylic substituted position to form para or ortho aQMs, respectively. With these constructs, hydroxylamine-based aQMs were generated and examined for the first time. Formation of the predicted intermediates was confirmed via trapping with Diels-Alder chemistry between an ortho aQM and furan. We also demonstrate aQM alkylation with nucleophiles such as thiols and phenzylhydrazine. While strong nucleophiles yield detectable adducts, weaker nucleophiles were unable to form adducts with these aQMs. Additionally, a para aQM alkylated a broader range of nucleophiles compared to an equivalent ortho aQM. These studies are motivated by our goal of designing nitroaromatic prodrugs that form aQMs for alkylation of essential nucleophiles such as proteins, nucleic acids, and other biomolecules.

Nitroreductase-dependent generation of aza-quinone methides for alkylation of biological nucleophiles

ACS Spring 2022

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