Bioinformatics-Guided Discovery of Citrulline 4-Hydroxylase from GE81112 Biosynthesis and Rational Engineering of Its Substrate Specificity

Abstract

We functionally characterize a nonheme dioxygenase from GE81112 biosynthesis and identify it as a citrulline hydroxylase. A bioinformatics guided engineering was performed to alter the substrate specificity of the enzyme.

Modular Chemoenzymatic Synthesis of GE81112 B1 and Related Analogues Enables Elucidation of Its Key Pharmacophores

“…We envisioned utilizing C–H hydroxylation reactions present in GE81112 biosynthesis to construct AA1 and AA2 (Figure B). Previous reports established GetF and GetI as iron and α-ketoglutarate dependent dioxygenase enzymes (Fe/αKG) responsible for hydroxylation of l -pipecolic acid ( l -Pip) and l -citrulline ( l -Cit), respectively (Figure B) . Next, a chemoselective azo coupling would establish a “masked” 2-aminoimidazole motif present within AA3, aiding purification and imparting chemoselectivity in subsequent transformations .…”