Structural Studies of the Stable Tyrosine Radical, YD + in Photosystem II

Barry, Bridgette A.; El-Deeb, M. K.; Sithole, I.; Debus, Richard J.; McIntosh, Lee; Babcock, Gerald T.

doi:10.1007/978-94-009-0511-5_109

Cited by 10 publications

(21 citation statements)

References 16 publications

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“…In fact two tyrosine radicals have been detected, 19,362 and their positions identified by sitedirected mutagenesis: Y Z • at position 161 of D 1 , 363,364 and Y D • at position 160 in D 2 . 365,366 Y Z • is directly involved in oxygen evolution and exists only transiently.…”

Section: B Proposed Roles Of Tyrosyl Radicals Y Z • and Y D •mentioning

confidence: 98%

“…Since the seminal experiments of Sjöberg et al identifying the organic radical essential for ribonucleotide reduction as being a one electron oxidized tyrosine residue, 15 stable and transient amino acid radicals have been localized to glycines, 16,17 cysteines, 18 tyrosines, 19,20 and tryptophans 21 as well as a variety of modified tyrosine 22,23 and tryptophan 24,25 residues within proteins. These species have been identified predominantly using electron paramagnetic resonance (EPR) spectroscopy of specifically isotopically labeled proteins.…”

Section: A One Electron Oxidized Amino Acids Thus Far Identified In mentioning

confidence: 99%

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Protein Radicals in Enzyme Catalysis

1998

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ContentsI. Introduction 705 II. General Principles 706 A. One Electron Oxidized Amino Acids Thus Far Identified in Proteins 706 B. Biosynthesis of (Modified) Amino Acid Radicals 706 C. Emerging General Chemical Properties of Enzymes Utilizing Protein Radicals for Catalysis 708 D. Methods To Examine Radical Dependent Reactions 710 III. Background on Ribonucleotide Reductases 710 IV. Class I Ribonucleotide Reductases 712 A. Formation of the Tyrosyl Radical 713 B. Function of the Tyrosyl Radical 714 C. Thiyl Radical Involvement in Catalysis 716 D. Evidence for Enzyme-Mediated Radical Chemistry Using Nucleotide Analogues 717 E. Structure 719 V. Class II Ribonucleotide Reductases 719 A. Exchange Reaction: Detection of the Elusive Thiyl Radical 719 B. Role of the Thiyl Radical in Catalysis 721 VI. Pyruvate Formate Lyase 722 A. Characterization of the Glycyl Radical 723 B. Formation of the Glycyl Radical 723 C. Catalytic Mechanism 724 D. Enzyme-Mediated Radical Chemistry with Pyruvate Analogues 727 VII. Anaerobic Ribonucleotide Reductase 728 A. Background 728 B. Requirement for a Glycyl Radical 728 C. Mechanism of Nucleotide Reduction 729 VIII. Cytochrome c Peroxidase 730 A. Formation of the Ferryl Heme/Tryptophan Cation Radical 730 B. Proposed Function of the Tryptophan Radical in Electron Transfer 731 IX. Prostaglandin H Synthase 733 A. Structure: A Tyrosine Residue Revealed 733 B. Proposed Role of the Tyrosyl Radical in Catalysis 734 X. Photosynthetic Oxygen Evolution 736 A. Background 736 B. Proposed Roles of Tyrosyl Radicals Y Z • and Y D • 737 C. Spectroscopic Studies on Y Z • and Y D • 738 D. Proposed Mechanisms for Water Oxidation and O 2 Evolution 739 XI. Galactose Oxidase 741 A. Different Redox States of Galactose Oxidase 741 B. Structure: A Modified Tyrosine Residue Revealed 742 C. Is the Oxidized Amino Acid Associated with Apo Oxidized GAO the Novel Ortho-Thiol-Substituted Tyrosine? 743 D. Catalytic Mechanism 743 XII. Quinoproteins 744 A. Copper-Dependent Amine Oxidases 744 B. Methylamine Dehydrogenase 749 XIII. Other Systems in Which Protein-Based Radicals Have Been Proposed or Detected 751 A. Bovine Liver Catalase 751 B. DNA Photolyase 751 C. Dopamine β Monooxygenase 752 XIV. Summary and Outlook 754 XV. Abbreviations 754 XVI. Acknowledgments 755 XVII. References 755 JoAnne Stubbe is the Novartis professor of Chemistry and Biology at the Massachusetts Institute of Technology. She received her undergraduate degree from the University of Pennsylvania working with Ed Thornton and did NSF-sponsored undergraduate research with Ed Trachtenberg at Clark University. These mentors played a strong role in her interest in physical organic chemistry. She received her Ph.D.

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Section: B Proposed Roles Of Tyrosyl Radicals Y Z • and Y D •mentioning

confidence: 98%

Section: A One Electron Oxidized Amino Acids Thus Far Identified In mentioning

confidence: 99%

Protein Radicals in Enzyme Catalysis

1998

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“…1 Of these, tyrosyl radicals (Y•s) have been shown to mediate a number of key metabolic transformations, including O 2 evolution, fatty acid oxidation, peroxide disproportionation, and prostaglandin synthesis. 2 The thermodynamics of Y oxidation require that proton transfer (PT) accompanies electron transfer (ET) at physiological pH, a process that may occur by either a stepwise or coupled (PCET) mechanism. To investigate the mechanism of Y• formation and Y•-mediated catalysis in metalloenzymes, a method by which the native Y• may be subtly perturbed is necessary.…”

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confidence: 99%

Incorporation of Fluorotyrosines into Ribonucleotide Reductase Using an Evolved, Polyspecific Aminoacyl-tRNA Synthetase

et al. 2011

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Tyrosyl radicals (Y•s) are prevalent in biological catalysis and are formed under physiological conditions by the coupled loss of both a proton and an electron. Fluorotyrosines (FnYs, n=1–4) are promising tools for studying the mechanism of Y• formation and reactivity, as their pKas and peak potentials span four units and 300 mV, respectively, between pH 6–10. In this manuscript, we present the directed evolution of aminoacyl-tRNA synthetases (aaRS) for 2,3,5-trifluorotyrosine (2,3,5-F3Y) and demonstrate their ability to charge an orthogonal tRNA with a series of FnYs, while maintaining high specificity over Y. An evolved aaRS is then used to site-specifically incorporate FnYs into the two subunits (α2 and β2) of E. coli class Ia ribonucleotide reductase (RNR), an enzyme that employs stable and transient Y•s to mediate long-range, reversible radical hopping during catalysis. Each of four conserved Ys in RNR is replaced with FnY(s) and the resulting proteins isolated in good yields. FnYs incorporated at position 122 of β2, the site of a stable Y• in the wt RNR, generate long-lived FnY•s that are characterized by EPR spectroscopy. Furthermore, we demonstrate that the radical pathway in the mutant Y122(2,3,5)F3Y-β2 is energetically and/or conformationally modulated such that the enzyme retains its activity, but that a new on-pathway Y• can accumulate. The distinct EPR properties of the 2,3,5-F3Y• facilitate spectral subtractions that make detection and identification of new Y•s straightforward.

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“…Thus Yi is reduced in microseconds by the oxygen evolving Mn, cluster and oxidized in nanoseconds by the reaction center PegO+; in contrast Y& is a stable radical, not essential for photosystem II activity, but somehow fundamental for the correct functioning of Yg [37]. The radicals are spectroscopically almost identical [38], but Yi has an increased relaxation rate, probably due to the proximity to the manganese site [35,36]. The tyrosines are buried in the middle of the membrane-spanning enzyme, and the phenol proton is not readily exchangeable.…”

Section: Photosystem IImentioning

confidence: 99%

“…Several of these enzymes use tyrosyl radicals with approximately the same spin density pattern [21,38,45,49], but their reactivity differs widely, depending on the protein environment [50]. The tryptophyl radical is clearly analogous, and TTQ and TOPA are modifications of the two amino acids; subdivision of these enzyme types is not reasonable.…”

Section: A Heterogeneous Family or An In-formal Gathering?mentioning

confidence: 99%

Protein‐radical enzymes

Pedersen

Finazzi‐Agrò

1993

FEBS Letters

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Protein-radical enzymes use a free radical located on an intrinsic amino acid residue as a cofactor. The amino acid involved can be a tyrosine (ribonucleotide reductase, photosystem II, prostaglandin H synthase), a modified tyrosine (amine oxidase, galactose oxidase), a tryptophan (cytochrome c peroxidase), a modified tryptophan (methylamine dehydrogenase) or a glycine (ribonucleotide reductase, pyruvate formate lyase). The mechanistic role of these radicals appears to be that of a one-electron gate, allowing the separation of single reducing equivalents in time and space.

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Structural Studies of the Stable Tyrosine Radical, YD + in Photosystem II

Cited by 10 publications

References 16 publications

Protein Radicals in Enzyme Catalysis

Protein Radicals in Enzyme Catalysis

Incorporation of Fluorotyrosines into Ribonucleotide Reductase Using an Evolved, Polyspecific Aminoacyl-tRNA Synthetase

Protein‐radical enzymes

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