UV-visible absorption spectrum of FAD and its reduced forms embedded in a cryptochrome protein

Schwinn, Karno; Ferré, Nicolas; Huix‐Rotllant, Miquel

doi:10.1039/d0cp01714k

Cited by 48 publications

(73 citation statements)

References 70 publications

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“…The UV-Vis spectrum of TmEnc has peaks at 360 and 450 nm, consistent with the spectra of flavins [Supplementary Fig. S1(c)] (Schwinn et al, 2020). Although the exact identity of the flavin ligand is ambiguous, flavin mononucleotide (FMN) seems to be consistent with the density and has been modeled into the structure.…”

Section: Flavin Ligandsupporting

confidence: 59%

Cryo-EM structure of a thermostable bacterial nanocompartment

Wiryaman

Toor

2021

IUCrJ

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Protein nanocompartments are widespread in bacteria and archaea, but their functions are not yet well understood. Here, the cryo-EM structure of a nanocompartment from the thermophilic bacterium Thermotoga maritima is reported at 2.0 Å resolution. The high resolution of this structure shows that interactions in the E-loop domain may be important for the thermostability of the nanocompartment assembly. Also, the channels at the fivefold axis, threefold axis and dimer interface are assessed for their ability to transport iron. Finally, an unexpected flavin ligand was identified on the exterior of the shell, indicating that this nanocompartment may also play a direct role in iron metabolism.

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Section: Flavin Ligandsupporting

confidence: 59%

Cryo-EM structure of a thermostable bacterial nanocompartment

Wiryaman

Toor

2021

IUCrJ

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“…According to Lactobacillus metabolism, these conditions lead to the production of the reduced forms of nicotinamide adenine dinucleotide (NADH) or nicotinamide adenine dinucleotide phosphate (NADPH), which, in absence of oxygen, lower the oxidation-reduction potential of the medium. This clue suggested to us the possibility that a reduced form of FAD [50], namely FAD •− , FADH • , FADH − , and FADH 2 , could be the actual activating agent of the enzyme.…”

Section: Introductionmentioning

confidence: 92%

Oleate Hydratase from Lactobacillus rhamnosus ATCC 53103: A FADH2-Dependent Enzyme with Remarkable Industrial Potential

Serra¹,

Simeis²,

Marzorati³

et al. 2021

Catalysts

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Recently, we described the preparation of the recombinant oleate hydratase from Lactobacillus rhamnosus ATCC 53103. We observed that the purified C-terminal His-tagged enzyme was completely inactive and the catalytic activity was partially restored only in presence of a large amount of flavin adenine dinucleotide (FAD). In the present work, we assess that this hydratase in the presence of the reduced form of flavin adenine dinucleotide (FADH2) is at least one hundred times as active as in the presence of the same concentration of FAD. By means of two different biochemical processes, we demonstrated unambiguously that oleate hydratase from Lactobacillus rhamnosus ATCC 53103 is a FADH2-dependent enzyme. As a first relevant application of this discovery, we devised a preparative procedure for the stereoselective synthesis of (R)-10-hydroxystearic acid. Accordingly, the hydration of oleic acid (up to 50 g/L) is performed on a multigram scale using the recombinant hydratase and FADH2 generated in situ as cofactor. The produced (R)-10-hydroxystearic acid (ee > 97%) precipitates from the reaction solvent (water/glycerol/ethanol) and is conveniently recovered by simple filtration (>90% yield).

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“…Upon photoillumination with light at 455 nm wavelength, these bands bleach and new absorption bands form at 350-400 nm. This spectral shape is characteristic of fully reduced FADH À (Kao et al, 2008;Schwinn et al, 2020). This reduction requires that two photons are absorbed and we therefore infer that the semiquinone (FAD À ) is formed as an intermediate.…”

Section: Resultsmentioning

confidence: 77%

The three-dimensional structure of Drosophila melanogaster (6–4) photolyase at room temperature

Cellini

Wahlgren

Henry

et al. 2021

Acta Cryst Sect D Struct Biol

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(6–4) photolyases are flavoproteins that belong to the photolyase/cryptochrome family. Their function is to repair DNA lesions using visible light. Here, crystal structures of Drosophila melanogaster (6–4) photolyase [Dm(6–4)photolyase] at room and cryogenic temperatures are reported. The room-temperature structure was solved to 2.27 Å resolution and was obtained by serial femtosecond crystallography (SFX) using an X-ray free-electron laser. The crystallization and preparation conditions are also reported. The cryogenic structure was solved to 1.79 Å resolution using conventional X-ray crystallography. The structures agree with each other, indicating that the structural information obtained from crystallography at cryogenic temperature also applies at room temperature. Furthermore, UV–Vis absorption spectroscopy confirms that Dm(6–4)photolyase is photoactive in the crystals, giving a green light to time-resolved SFX studies on the protein, which can reveal the structural mechanism of the photoactivated protein in DNA repair.

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UV-visible absorption spectrum of FAD and its reduced forms embedded in a cryptochrome protein

Abstract: Simulation of UV-vis absorption spectra of cryptochromes and flavoproteins requires an explicit account of vibrations of the flavin chromophore embedded in protein.

Cited by 48 publications

References 70 publications

Cryo-EM structure of a thermostable bacterial nanocompartment

Cryo-EM structure of a thermostable bacterial nanocompartment

Oleate Hydratase from Lactobacillus rhamnosus ATCC 53103: A FADH2-Dependent Enzyme with Remarkable Industrial Potential

The three-dimensional structure of Drosophila melanogaster (6–4) photolyase at room temperature

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