Nature uses highly reactive radicals to carry out a diverse set of biochemical functions, many of which are essen- tial to maintaining proper human health. These potent biological radical reactions need to be carried out safely, producing essential specific products, without dangerous side reactions occurring. A large number of such rad- ical reactions are performed by the family of radical SAM enzymes, which use a [4Fe-4S] center with a bound S-adenosylmethionine (SAM) molecule to generate a strongly oxidizing 5'-deoxyadenosyl radical which can in turn drive a large number of difficult chemical reactions. We will target mechanistic aspects of several classes of radical SAM enzymes. Biotin synthase is a radical SAM enzyme that catalyzes the final step in the biosynthesis of the vitamin biotin. A set of Fe-S and radical SAM maturase enzymes are used to build the unique Fe-S center of Fe-Fe hydrogenase, an enzyme which catalyzes the important reduction of protons to dihydrogen and vice versa. And radical SAM enzymes are used to modify many bases in transfer RNA, improving codon-anticodon recognition in order to make protein synthesis more reliable. We are specifically interested in a radical SAM enzyme QueE that is essential for generating 7-deazapurines. This proposal describes a magnetic resonance spectroscopic approach to study such diverse radical SAM enzymes. Specifically, we are using electron para- magnetic resonance (EPR) spectroscopy, which can precisely measure the magnetic environment of unpaired electrons in the radical SAM Fe-S clusters, in the organic radicals that these clusters generate, and in secondary metal centers that are involved in the reactions in many of these enzymes.
Radicals, high-energy chemical species with unpaired electrons, can cause deleterious reactions in biology, including those adversely affecting human health. At the same time the potent reactions radicals can carry out are necessary for a large number of crucial enzymes. We are studying a set of these radical reactions to learn how biological metal centers and radicals work in concert to safely harness the power of radical reaction chemistry in the superfamily of Radical SAM Enzymes.
|Barr, Ian; Stich, Troy A; Gizzi, Anthony S et al. (2018) X-ray and EPR Characterization of the Auxiliary Fe-S Clusters in the Radical SAM Enzyme PqqE. Biochemistry 57:1306-1315|
|Hadley, Rose C; Gagnon, Derek M; Brophy, Megan Brunjes et al. (2018) Biochemical and Spectroscopic Observation of Mn(II) Sequestration from Bacterial Mn(II) Transport Machinery by Calprotectin. J Am Chem Soc 140:110-113|
|Guo, Yirui; Suess, Daniel L M; Herzik Jr, Mark A et al. (2017) Regulation of nitric oxide signaling by formation of a distal receptor-ligand complex. Nat Chem Biol 13:1216-1221|
|Bruender, Nathan A; Wilcoxen, Jarett; Britt, R David et al. (2016) Biochemical and Spectroscopic Characterization of a Radical S-Adenosyl-L-methionine Enzyme Involved in the Formation of a Peptide Thioether Cross-Link. Biochemistry 55:2122-34|
|Suess, Daniel L M; Kuchenreuther, Jon M; De La Paz, Liliana et al. (2016) Biosynthesis of the [FeFe] Hydrogenase H Cluster: A Central Role for the Radical SAM Enzyme HydG. Inorg Chem 55:478-87|
|Wilcoxen, Jarett; Arragain, Simon; Scandurra, Alessandro A et al. (2016) Electron Paramagnetic Resonance Characterization of Three Iron-Sulfur Clusters Present in the Nitrogenase Cofactor Maturase NifB from Methanocaldococcus infernus. J Am Chem Soc 138:7468-71|
|Zhu, Wen; Wilcoxen, Jarett; Britt, R David et al. (2016) Formation of Hexacoordinate Mn(III) in Bacillus subtilis Oxalate Decarboxylase Requires Catalytic Turnover. Biochemistry 55:429-34|
|Suess, Daniel L M; Pham, Cindy C; Bürstel, Ingmar et al. (2016) The Radical SAM Enzyme HydG Requires Cysteine and a Dangler Iron for Generating an Organometallic Precursor to the [FeFe]-Hydrogenase H-Cluster. J Am Chem Soc 138:1146-9|
|Suess, Daniel L M; Bürstel, Ingmar; De La Paz, Liliana et al. (2015) Cysteine as a ligand platform in the biosynthesis of the FeFe hydrogenase H cluster. Proc Natl Acad Sci U S A 112:11455-60|
|Miles, Zachary D; Myers, William K; Kincannon, William M et al. (2015) Biochemical and Spectroscopic Studies of Epoxyqueuosine Reductase: A Novel Iron-Sulfur Cluster- and Cobalamin-Containing Protein Involved in the Biosynthesis of Queuosine. Biochemistry 54:4927-35|
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