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Publication Abstract from PubMed

Rieske oxygenases exploit the reactivity of iron to perform chemically challenging C-H bond functionalization reactions. Thus far, only a handful of Rieske oxygenases have been structurally characterized and remarkably little information exists regarding how these enzymes use a common architecture and set of metallocenters to facilitate a diverse range of reactions. Herein, we detail how two Rieske oxygenases SxtT and GxtA use different protein regions to influence the site-selectivity of their catalyzed monohydroxylation reactions. We present high resolution crystal structures of SxtT and GxtA with the native beta-saxitoxinol and saxitoxin substrates bound in addition to a Xenon-pressurized structure of GxtA that reveals the location of a substrate access tunnel to the active site. Ultimately, this structural information allowed for the identification of six residues distributed between three regions of SxtT that together control the selectivity of the C-H hydroxylation event. Substitution of these residues produces a SxtT variant that is fully adapted to exhibit the non-native site-selectivity and substrate scope of GxtA. Importantly, we also found that these selectivity regions are conserved in other structurally characterized Rieske oxygenases, providing a framework for predictively repurposing and manipulating Rieske oxygenases as biocatalysts.

Design principles for site-selective hydroxylation by a Rieske oxygenase.,Liu J, Tian J, Perry C, Lukowski AL, Doukov TI, Narayan ARH, Bridwell-Rabb J Nat Commun. 2022 Jan 11;13(1):255. doi: 10.1038/s41467-021-27822-3. PMID:35017498^[1]

From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine.