title: Diradical intermediate within the context of tryptophan tryptophylquinone biosynthesis.
abstract: Despite the importance of tryptophan (Trp) radicals in biology, very few radicals have been trapped and characterized in a physiologically meaningful context. Here we demonstrate that the diheme enzyme MauG uses Trp radical chemistry to catalyze formation of a Trp-derived tryptophan tryptophylquinone cofactor on its substrate protein, premethylamine dehydrogenase. The unusual six-electron oxidation that results in tryptophan tryptophylquinone formation occurs in three discrete two-electron catalytic steps. Here the exact order of these oxidation steps in the processive six-electron biosynthetic reaction is determined, and reaction intermediates are structurally characterized. The intermediates observed in crystal structures are also verified in solution using mass spectrometry. Furthermore, an unprecedented Trp-derived diradical species on premethylamine dehydrogenase, which is an intermediate in the first two-electron step, is characterized using high-frequency and -field electron paramagnetic resonance spectroscopy and UV-visible absorbance spectroscopy. This work defines a unique mechanism for radical-mediated catalysis of a protein substrate, and has broad implications in the areas of applied biocatalysis and understanding of oxidative protein modification during oxidative stress.
authors.affiliation: Department of Biochemistry, Molecular Biology and Biophysics, University of Minnesota, Minneapolis, MN 55455, USA.
authors.forename: Erik T
authors.initials: ET
authors.lastname: Yukl
authors.forename: Fange
authors.initials: F
authors.lastname: Liu
authors.forename: J
authors.initials: J
authors.lastname: Krzystek
authors.forename: Sooim
authors.initials: S
authors.lastname: Shin
authors.forename: Lyndal M R
authors.initials: LM
authors.lastname: Jensen
authors.forename: Victor L
authors.initials: VL
authors.lastname: Davidson
authors.forename: Carrie M
authors.initials: CM
authors.lastname: Wilmot
authors.forename: Aimin
authors.initials: A
authors.lastname: Liu
mesh_terms: D001426:Bacterial Proteins / Q000737:chemistry* / Q000378:metabolism; D018360:Crystallography, X-Ray; D045563:Indolequinones / Q000096:biosynthesis* / Q000737:chemistry; D013058:Mass Spectrometry; D010084:Oxidation-Reduction; D010088:Oxidoreductases / Q000737:chemistry* / Q000378:metabolism; D010231:Paracoccus denitrificans / Q000201:enzymology*; D013379:Substrate Specificity; D014364:Tryptophan / Q000031:analogs & derivatives* / Q000096:biosynthesis / Q000737:chemistry
chemical_list: D001426:Bacterial Proteins; D045563:Indolequinones; C070569:tryptophan tryptophylquinone; D014364:Tryptophan; D010088:Oxidoreductases