Crystal Structures of the Lumazine Protein from<i>Photobacterium kishitanii</i>in Complexes with the Authentic Chromophore, 6,7-Dimethyl- 8-(1′-<scp>d</scp>-Ribityl) Lumazine, and Its Analogues, Riboflavin and Flavin Mononucleotide, at High Resolution

Sato, Yuichi; Shimizu, Satoshi; Ohtaki, Akashi; Noguchi, Keiichi; Miyatake, Hideyuki; Dohmae, Naoshi; Sasaki, Satoshi; Odaka, Masafumi; Yohda, Masafumi

doi:10.1128/jb.01015-09

Cited by 26 publications

(16 citation statements)

References 41 publications

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“…The fluorescence is readily quenched by KI meaning that the ligand is bound to a solvent‐exposed site as now verified in the spatial structures (Figs. and ) . The interactions of the ligand within the binding cavity are consistent with the conclusions from NMR .…”

Section: Introductionsupporting

confidence: 82%

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The Sensitized Bioluminescence Mechanism of Bacterial Luciferase

Lee

Müller²,

Visser

2018

Photochem & Photobiology

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After more than one‐half century of investigations, the mechanism of bioluminescence from the FMNH2 assisted oxygen oxidation of an aliphatic aldehyde on bacterial luciferase continues to resist elucidation. There are many types of luciferase from species of bioluminescent bacteria originating from both marine and terrestrial habitats. The luciferases all have close sequence homology, and in vitro, a highly efficient light generation is obtained from these natural metabolites as substrates. Sufficient exothermicity equivalent to the energy of a blue photon is available in the chemical oxidation of the aldehyde to the corresponding carboxylic acid, and a luciferase‐bound FMNH‐OOH is a key player. A high energy species, the source of the exothermicity, is unknown except that it is not a luciferin cyclic peroxide, a dioxetanone, as identified in the pathway of the firefly and the marine bioluminescence systems. Besides these natural substrates, variable bioluminescence properties are found using other reactants such as flavin analogs or aldehydes, but results also depend on the luciferase type. Some rationalization of the mechanism has resulted from spatial structure determination, NMR of intermediates and dynamic optical spectroscopy. The overall light path appears to fall into the sensitized class of chemiluminescence mechanism, distinct from the dioxetanone types.

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Section: Introductionsupporting

confidence: 82%

“…The spatial structure of LumP modeled with an associated Lux is consistent with this separation (Fig. ) .…”

Section: Introductionsupporting

confidence: 71%

The Sensitized Bioluminescence Mechanism of Bacterial Luciferase

Lee

Müller²,

Visser

2018

Photochem & Photobiology

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“…15,16,41,42 Recently, on the basis of the crystal structures, a computational model of the luciferaseÀlumazine protein complex has been constructed. 43 It reveals a donorÀacceptor separation of <20 Å.…”

Section: ' Discussionmentioning

confidence: 99%

Green-Fluorescent Protein from the Bioluminescent Jellyfish Clytia gregaria Is an Obligate Dimer and Does Not Form a Stable Complex with the Ca²⁺-Discharged Photoprotein Clytin

Malikova

Visser²,

Hoek³

et al. 2011

Biochemistry

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Green-fluorescent protein (GFP) is the origin of the green bioluminescence color exhibited by several marine hydrozoans and anthozoans. The mechanism is believed to be Förster resonance energy transfer (FRET) within a luciferase-GFP or photoprotein-GFP complex. As the effect is found in vitro at micromolar concentrations, for FRET to occur this complex must have an affinity in the micromolar range. We present here a fluorescence dynamics investigation of the recombinant bioluminescence proteins from the jellyfish Clytia gregaria, the photoprotein clytin in its Ca(2+)-discharged form that is highly fluorescent (λ(max) = 506 nm) and its GFP (cgreGFP; λ(max) = 500 nm). Ca(2+)-discharged clytin shows a predominant fluorescence lifetime of 5.7 ns, which is assigned to the final emitting state of the bioluminescence reaction product, coelenteramide anion, and a fluorescence anisotropy decay or rotational correlation time of 12 ns (20 °C), consistent with tight binding and rotation with the whole protein. A 34 ns correlation time combined with a translational diffusion constant and molecular brightness from fluorescence fluctuation spectroscopy all confirm that cgreGFP is an obligate dimer down to nanomolar concentrations. Within the dimer, the two chromophores have a coupled excited-state transition yielding fluorescence depolarization via FRET with a transfer correlation time of 0.5 ns. The 34 ns time of cgreGFP showed no change upon addition of a 1000-fold excess of Ca(2+)-discharged clytin, indicating no stable complexation below 0.2 mM. It is proposed that any bioluminescence FRET complex with micromolar affinity must be one formed transiently by the cgreGFP dimer with a short-lived (millisecond) intermediate in the clytin reaction pathway.

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“…Recently, studies on the crystal structure of the lumazine protein complexed with riboflavin and 6,7-dimethyl-8-ribityllumazine have been published. 11,12 They show that in the ligand-binding site an extensive direct and watermediated hydrogen bond network is present. Apparently these intermolecular interactions are required for the correct function of the lumazine cofactor.…”

Section: Introductionmentioning

confidence: 98%

Vibrational Spectra of the Ground and the Singlet Excited ππ* State of 6,7-Dimethyl-8-ribityllumazine

Schreier¹,

Pugliesi²,

Koller³

et al. 2011

J. Phys. Chem. B

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6,7-Dimethyl-8-ribityllumazine serves as fluorophore in lumazine proteins (LumP) of luminescent bacteria. The molecule exhibits several characteristic vibrational absorption bands between 1300 and 1750 cm(-1) in its electronic ground state. The IR-absorption pattern of the singlet excited ππ* state was monitored via ultrafast infrared spectroscopy after photoexcitation at 404 nm. The comparison of experimentally observed band shifts for a number of isotopologues allows for a clear assignment of several absorption bands--most importantly the two carbonyl bands. This assignment is confirmed by normal-mode calculations by means of either density functional theory (DFT) calculations for the ground state or the configuration interaction singles (CIS) method for the excited singlet state. A good agreement between experiment and calculation is obtained for models including explicitly a first solvation shell. The results provide a basis for further investigations of lumazine protein and demonstrate the necessity of proper accounting for explicit hydrogen bonding in case of strongly polar molecular systems.

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Crystal Structures of the Lumazine Protein fromPhotobacterium kishitaniiin Complexes with the Authentic Chromophore, 6,7-Dimethyl- 8-(1′-d-Ribityl) Lumazine, and Its Analogues, Riboflavin and Flavin Mononucleotide, at High Resolution

Cited by 26 publications

References 41 publications

The Sensitized Bioluminescence Mechanism of Bacterial Luciferase

The Sensitized Bioluminescence Mechanism of Bacterial Luciferase

Green-Fluorescent Protein from the Bioluminescent Jellyfish Clytia gregaria Is an Obligate Dimer and Does Not Form a Stable Complex with the Ca²⁺-Discharged Photoprotein Clytin

Vibrational Spectra of the Ground and the Singlet Excited ππ* State of 6,7-Dimethyl-8-ribityllumazine

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Crystal Structures of the Lumazine Protein fromPhotobacterium kishitaniiin Complexes with the Authentic Chromophore, 6,7-Dimethyl- 8-(1′-d-Ribityl) Lumazine, and Its Analogues, Riboflavin and Flavin Mononucleotide, at High Resolution

Cited by 26 publications

References 41 publications

The Sensitized Bioluminescence Mechanism of Bacterial Luciferase

The Sensitized Bioluminescence Mechanism of Bacterial Luciferase

Green-Fluorescent Protein from the Bioluminescent Jellyfish Clytia gregaria Is an Obligate Dimer and Does Not Form a Stable Complex with the Ca2+-Discharged Photoprotein Clytin

Vibrational Spectra of the Ground and the Singlet Excited ππ* State of 6,7-Dimethyl-8-ribityllumazine

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Product

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Green-Fluorescent Protein from the Bioluminescent Jellyfish Clytia gregaria Is an Obligate Dimer and Does Not Form a Stable Complex with the Ca²⁺-Discharged Photoprotein Clytin