New roles of flavoproteins in molecular cell biology: Blue‐light active flavoproteins studied by electron paramagnetic resonance

Schleicher, Erik; Bittl, Robert; Weber, Stefan

doi:10.1111/j.1742-4658.2009.07141.x

Cited by 23 publications

(26 citation statements)

References 93 publications

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“…All the detected spectral features correspond to protons within the flavin ring in the weakcoupling regime, which are easy to assign from previously reported measurements. 25,26 The region containing the matrix peak and very weakly interacting protons (solvent and protein protons together with H3, CH 3 (7) and H9 of the flavin ring) is marked with the label a in Figure 2. Signals assigned to flavin proton H6, labelled as b, CH 3 (8), labelled as c, and H5, labelled as f, yield hyperfine constants (see Table 1) fully coherent with those previously reported for the same protein.…”

Section: 1-experimental Resultsmentioning

confidence: 99%

Methyl rotors in flavoproteins

Martínez

Alonso

García-Rubio

et al. 2014

Phys. Chem. Chem. Phys.

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In this contribution we present the study of the thermal dependence of the ENDOR spectra of flavodoxin at low temperatures which reveals the dynamics of the methyl groups bound to the flavin moiety in flavoproteins. The methyl groups behave as quantum rotors locked by a deep rotational well and undergoing a tunneling process. At room temperature, methyl rotors are locked and the hopping motion is slow. This picture of the dynamics of the methyl groups of the flavin ring is quite different from the one usually accepted and has relevant consequences on the understanding of the mechanisms of flavoproteins.

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Section: 1-experimental Resultsmentioning

confidence: 99%

Methyl rotors in flavoproteins

Martínez

Alonso

García-Rubio

et al. 2014

Phys. Chem. Chem. Phys.

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“…The g tensor is rhombic, and with the help of the hydrogen standard the principal values were determined as 2.00430(5), 2.00353(5) and 2.00210(9), values very similar to those of anionic flavin radicals (e.g. [58; 59]). From these accurate g values, the protonation state of the cofactor can be determined [60].…”

Section: Examplesmentioning

confidence: 99%

Atomic hydrogen as high-precision field standard for high-field EPR

Stoll

Ozarowski

Britt

et al. 2010

Journal of Magnetic Resonance

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We introduce atomic hydrogen trapped in an octaisobutylsilsesquioxane nanocage (H@iBuT 8 ) as a new molecular high-precision magnetic field standard for high-field EPR spectroscopy of organic radicals and other systems with signals around g = 2. Its solid-state EPR spectrum consists of two narrow lines separated by about 51 mT and centered at g ≈ 2. The isotropic g factor is 2.00294(3) and essentially temperature independent. The isotopic 1 H hyperfine coupling constant is 1416.8(2) MHz below 70 K and decreases slightly with increasing temperature to 1413.7(1) MHz at room temperature. The spectrum of the standard does not overlap with those of most organic radicals, and it can be easily prepared and is stable at room temperature.

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“…[19,20] The formation of tyrosyl radicals is known to occur in other flavoproteins, for example, photolyases and cryptochromes. [21][22][23] The presence of a neutral flavosemiquinone and a neutral tyrosine radical has also recently been proposed to exist in the blue light sensor protein TePixD. [24] However, unlike the other known examples, in which radical formation follows photoactivation of the flavin, MAO is novel in that it is the ground state semiquinone form of the FAD that establishes the redox equilibrium with the amino acid radical species.…”

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Tyrosyl Radical Formation and Propagation in Flavin Dependent Monoamine Oxidases

et al. 2010

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Understanding the mechanism of biological amine oxidation by flavoproteins remains a controversial area of research. This is particularly true for the mammalian monoamine oxidases (MAO), which are major pharmaceutical targets for the development of antidepressants and neuroprotective agents.[1] Despite intensive research efforts, the detailed reaction mechanism, which will have major implications for drug discovery with this class of enzyme, has yet to be discovered. Mechanistic debates have centred on the potential involvement of radical species in the activity of these enzymes. We now provide strong evidence that supports a role for radical catalysis in flavoprotein monoamine oxidases.The mammalian monoamine oxidases (EC 1.4.3.4) are localised to the outer mitochondrial membrane. In these enzymes, the flavin cofactor, FAD, is covalently linked through the 8a-methyl group to an active site cysteine residue.[2] A number of mechanisms for MAO-catalysed amine oxidation have been proposed over the years, and several reviews are available. [3][4][5][6] There are currently three main mechanistic proposals for MAO catalysis. These comprise: 1) the concerted polar nucleophilic mechanism; 2) the direct hydride transfer mechanism; and 3) the single electron transfer mechanism. Recent support for the concerted polar nucleophilic mechanism has come from kinetic and structural studies on tyrosine mutants of MAO B, [7] and also from computational studies. [8,9] However, analysis of the nitrogen isotope effects conducted on a related amine oxidase, Nmethyltryptophan oxidase, supported either a direct hydride transfer mechanism or, possibly, a discrete electron transfer mechanism.[10] The single electron transfer (SET) mechanism initially proposed by Silverman and colleagues involves single electron transfer from the substrate nitrogen lone pair to yield the substrate radical and flavin semiquinone.[11] The SET mechanism is consistent with electronic effects observed in quantitative structure activity relationships studies with a series of substituted benzylamines, but the identity of the one-electron oxidant required for the formation of the aminyl radical cation was a major concern. [12] Recent spectroscopic evidence for the presence of a stable tyrosyl radical in partially reduced human MAO A provided support for the SET mechanism and led to the proposal of a modified SET mechanism (Scheme 1).[13] After the initial singleelectron transfer from the amine to the flavin to form the aminyl radical cation and flavin semiquinone, a redox equilibrium was proposed between the semiquinone and a neutral tyrosyl radical(s) during the catalytic cycle. On initial inspection, it might seem thermodynamically unfeasible that a ground state flavin (E m~À 0.2 to 0 V) is capable of oxidising a tyrosine residue (E m~0 .9 V). However, large apparent barriers to electron transfer might not prevent the reaction in an enzyme active site. For example, endergonic tunnelling over a relatively short distance followed by a thermodynamically favourable s...

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New roles of flavoproteins in molecular cell biology: Blue‐light active flavoproteins studied by electron paramagnetic resonance

Cited by 23 publications

References 93 publications

Methyl rotors in flavoproteins

Methyl rotors in flavoproteins

Atomic hydrogen as high-precision field standard for high-field EPR

Tyrosyl Radical Formation and Propagation in Flavin Dependent Monoamine Oxidases

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