(2jd) Probing the Mechanism of Isonitrile Formation By a Non-Heme Iron(II)-Dependent Oxidase/Decarboxylase

Teaching Interests

The isonitrile moiety is an electron-rich functionality that decorates various bioactive natural products isolated from diverse kingdoms of life. Isonitrile biosynthesis was restricted for over a decade to isonitrile synthases, a family of enzymes catalyzing a condensation reaction between ʟ-Trp/ʟ-Tyr and ribulose-5-phosphate. The discovery of ScoE, a non-heme iron(II) and α-ketoglutarate-dependent dioxygenase, demonstrated an alternative pathway employed by nature for isonitrile installation. Biochemical, crystallographic, and computational investigations of ScoE have previously been reported, yet the isonitrile formation mechanism remains obscure. In this present work, we employed in vitro biochemistry, chemical synthesis, spectroscopy techniques, and computational simulations that enabled us to propose a plausible molecular mechanism for isonitrile formation. Our findings demonstrate that the ScoE reaction initiates with C5 hydroxylation of (R)-3-((carboxymethyl)amino)butanoic acid to generate C5-OH CABA, which undergoes dehydration presumably mediated by Tyr96 to synthesize imine-CABA in a trans configuration. (R)-3-isocyanobutanoic acid is finally generated through radical-based decarboxylation of trans-imine-CABA, instead of the common hydroxylation pathway employed by this enzyme superfamily.