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Saleem-Batcha, R, Stull F, Sanders JN, Moore BS, Palfey BA, Houk KN, Teufel R.  2018.  Enzymatic control of dioxygen binding and functionalization of the flavin cofactor. Proceedings of the National Academy of Sciences of the United States of America. 115:4909-4914.   10.1073/pnas.1801189115   AbstractWebsite

The reactions of enzymes and cofactors with gaseous molecules such as dioxygen (O-2) are challenging to study and remain among the most contentious subjects in biochemistry. To date, it is largely enigmatic how enzymes control and fine-tune their reactions with O-2, as exemplified by the ubiquitous flavin-dependent enzymes that commonly facilitate redox chemistry such as the oxygenation of organic substrates. Here we employ O-2-pressurized X-ray crystallography and quantum mechanical calculations to reveal how the precise positioning of O-2 within a flavoenzyme's active site enables the regio-specific formation of a covalent flavin-oxygen adduct and oxygenating species (i.e., the flavin-N5-oxide) by mimicking a critical transition state. This study unambiguously demonstrates how enzymes may control the O-2 functionalization of an organic cofactor as prerequisite for oxidative catalysis. Our work thus illustrates how O-2 reactivity can be harnessed in an enzymatic environment and provides crucial knowledge for future rational design of O-2-reactive enzymes.

Jordan, PA, Moore BS.  2016.  Biosynthetic pathway connects cryptic ribosomally synthesized posttranslationally modified peptide genes with pyrroloquinoline alkaloids. Cell Chemical Biology. 23:1504-1514.   10.1016/j.chembiol.2016.10.009   AbstractWebsite

In an era where natural product biosynthetic gene clusters can be rapidly identified from sequenced genomes, it is unusual for the biosynthesis of an entire natural product class to remain unknown. Yet, the genetic determinates for pyrroloquinoline alkaloid biosynthesis have remained obscure despite their abundance and deceptive structural simplicity. In this work, we have identified the biosynthetic gene cluster for ammosamides A-C, pyrroloquinoline alkaloids from Streptomyces sp. CNR-698. Through direct cloning, heterologous expression and gene deletions we have validated the ammosamide biosynthetic gene cluster and demonstrated that these seemingly simple molecules are derived from a surprisingly complex set of biosynthetic genes that are also found in the biosynthesis of lymphostin, a structurally related pyrroloquinoline alkaloid from Salinispora and Streptomyces. Our results implicate a conserved set of genes driving pyrroloquinoline biosynthesis that consist of genes frequently associated with ribosomal peptide natural product biosynthesis, and whose exact biochemical role remains enigmatic.

Teufel, R, Miyanaga A, Michaudel Q, Stull F, Louie G, Noel JP, Baran PS, Palfey B, Moore BS.  2013.  Flavin-mediated dual oxidation controls an enzymatic Favorskii-type rearrangement. Nature. 503:552-+.   10.1038/nature12643   AbstractWebsite

Flavoproteins catalyse a diversity of fundamental redox reactions and are one of the most studied enzyme families(1,2). As monooxygenases, they are universally thought to control oxygenation by means of a peroxyflavin species that transfers a single atom of molecular oxygen to an organic substrate(1,3,4). Here we report that the bacterial flavoenzyme EncM(5,6) catalyses the peroxyflavin-independent oxygenation-dehydrogenation dual oxidation of a highly reactive poly(beta-carbonyl). The crystal structure of EncM with bound substrate mimics and isotope labelling studies reveal previously unknown flavin redox biochemistry. We show that EncM maintains an unexpected stable flavin-oxygenating species, proposed to be a flavin-N5-oxide, to promote substrate oxidation and trigger a rare Favorskii-type rearrangement that is central to the biosynthesis of the antibiotic enterocin. This work provides new insight into the fine-tuning of the flavin cofactor in offsetting the innate reactivity of a polyketide substrate to direct its efficient electrocyclization.