Light-Induced, Lysine-Targeting Irreversible Covalent Inhibition of the Human Oxygen Sensing Hydroxylase Factor Inhibiting HIF (FIH)

Factor inhibiting hypoxia-inducible factor (FIH) is a JmjC domain 2-oxoglutarate (2OG) and Fe(II)-dependent oxygenase that catalyzes protein hydroxylations, including of specific asparagines in the <i>C</i>-terminal transcriptional activation domains of hypoxia-inducible factor alpha (HIF-α) isoforms. FIH is of medicinal interest due to its ability to alter metabolism and modulate the course of the HIF-mediated hypoxic response. We report the development of a light-induced, lysine (Lys106)-targeting irreversible covalent inhibitor of FIH. The approach is complementary to optogenetic methods for regulation of transcription. The covalently reacting inhibitor <b>NBA-ZG-2291</b> was the result of structure-guided modification of the reported active site binding FIH inhibitor <b>ZG-2291</b> with an appropriately positioned <i>o</i>-nitrobenzyl alcohol (<i>o</i>-NBA) group. The results demonstrate that <b>NBA-ZG-2291</b> forms a stable covalent bond in a light-dependent process with Lys106 of FIH, inactivating its hydroxylation activity and resulting in sustained upregulation of FIH-dependent HIF target genes. The light-controlled inhibitors targeting a lysine residue enable light and spatiotemporal control of FIH activity in a manner useful for dissecting the context-dependent physiological roles of FIH.