Abstract 2471 Supercharged Heme: Unmasking a New Regime of Aromatic Oxidation by an Emergent Superfamily

Aromatic oxidation, a ubiquitous biological transformation, shapes the landscapes of natural products, pharmaceuticals, and environmental processes. For decades, heme-dependent enzymes like P450s and peroxygenases reigned supreme in this realm, wielding thiol-ligated hemes to insert oxygen into hydrocarbons. However, our recent discovery disrupts this established paradigm. We unveil a vast, previously unrecognized heme-dependent aromatic oxygenase (HDAO) superfamily,1 rewriting the playbook of aromatic oxidation with the power of histidine-ligated heme. Our quest to explore heme enzymes beyond the known led us to the TDO (tryptophan 2,3-dioxygenase) family, where we unearthed a hidden superfamily harboring immense functional diversity. Its more than eight distinct clades encompass thousands of members, including newly characterized MarE, SfmD, TyrH, and a captivating collection of emerging tryptophan hydroxylases (Arg5, Luz15, KerI, OhmK, and PsmG). These enzymes transcend the limitations previously ascribed to histidine-ligated hemes, wielding them to execute a dazzling array of C–H activation and functionalization, tackling a broader spectrum of aromatic substrates with exquisite precision. This presentation discusses our captivating journey as we delve deeper into the HDAO superfamily, exploring its novel reactions,2 unique cofactors,3 and vast substrate repertoire. We reveal unexpected aromatic transformations, including a novel C–F activation mediated by the intricate oxidizing power of the heme-activated oxidant in TyrH,4 through detailed structural analyses, biophysical characterization, and catalytic experimentation.2-4 The remarkable chemical prowess demonstrated by HDAO enzymes opens a new chapter in our understanding of aromatic transformations. This discovery holds significant potential for biocatalysis, drug discovery, and unlocking new bioinorganic and biological chemistry frontiers. References: 1) Shin I, Wang Y, and Liu A (2021) PNAS 2021, 118, e210656 (https://doi.org/10.1073/pnas.2106561118) 2) Wang Y, Davis I, Shin I, Wherritt DJ, Griffith WP, Dornevil K, Colabroy KL, and Liu A (2019) ACS Catalysis, 9, 4764-4776 (https://doi.org/10.1021/acscatal.9b00231) 3) Shin I, Davis I, Nieves-Merced K, Wang Y, McHardy S, and Liu A (2021) Chem. Sci., 12(11), 3984-3998 (https://doi.org/10.1039/D0SC06369J) 4) Wang Y, Davis I, Shin I, Xu H, and Liu A (2021) J. Am. Chem. Soc., 143, 4680-4693 (https://doi.org/10.1021/jacs.1c00175) This study of novel heme chemistry is funded by NIH grants R01GM108988, R21AG078775, R21CA270879, and Welch Foundation grant AX-2110-20220331.