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Spectral properties and function of 2 lumazine proteins from photobacterium


, : Spectral properties and function of 2 lumazine proteins from photobacterium. Biochemistry 24(6): 1476-1483

The spectral properties are compared for 2 6,7-dimethyl-8-ribityllumazine proteins from marine bioluminescent bacteria, 1 from a psychrophile, Photobacterium phosphoreum, and the other from a thermophile, Photobacterium leiognathi. The visible spectral properties, which are the ones by which the protein performs its biological function of bioluminescence emission, are almost the same for the 2 proteins: at 2.degree. C and 50 mM Pi, pH 7, fluorescence quantum yield .vphi.F = 0.59 and 0.54, respectively; fluorescence lifetime .tau. = 14.4 and 14.8 ns, respectively; fluorescence maxima, both 475 nm; absorption maximum, 417 and 420 nm, respectively; circular dichroism minima at around 420 nm, both -41 .times. 103 deg cm2 dmol-1. The ligand binding sites therefore must provide very similar environments and arguments are presented that the bound ligand is relatively exposed to solvent. The dissociation equilibrium was studied by steady-state fluorescence polarization. The thermophilic protein binds the ligand with Kd (20.degree. C) = 0.016 .mu.M, 10 times more tightly than the other protein [Kd (20.degree. C) = 0.16 .mu.M]. The origin of the binding difference probably resides in differences in secondary structure. The tryptophan fluorescence spectra of the 2 proteins are different, but more significant is an observation of the decay of the tryptophan emission anisotropy. For the psychrophilic lumazine protein this anisotropy decays to zero in 1 ns, implying that its single tryptophan residue lies in a very floppy region of the protein. For the other protein, the anisotropy exhibits both a fast component and a slow one corresponding to rotation of the protein as a whole. This suggests that in the thermophilic protein the tryptophan region is held more rigidly. In both proteins, however, the ligand exhibits no independent mobility, as its rotational correlation time (respectively 19.5 and 17.5 ns, 2.degree. C) corresponds to the rotation of a sphere of hydrated MW .apprx. 30,000.

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Related references

Lee, J.; O'Kane, D.J.; Visser, A.J., 1985: Spectral properties and function of two lumazine proteins from Photobacterium. The spectral properties are compared for two 6,7-dimethyl-8-ribityllumazine proteins from marine bioluminescent bacteria, one from a psychrophile, Photobacterium phosphoreum, and the other from a thermophile, Photobacterium leiognathi. The visible...

O'Kane, D.J.; Karle, V.A.; Lee, J., 1985: Purification of lumazine proteins from Photobacterium leiognathi and Photobacterium phosphoreum: bioluminescence properties. Bright strains of the marine bioluminescent bacterium Photobacterium leiognathi produce a "lumazine protein" in amounts comparable to that previously found in Photobacterium phosphoreum. New protocols are developed for the purification t...

O'Kane, D.J.; Lee, J., 1986: Purification and properties of lumazine proteins from Photobacterium strains. Methods in Enzymology 133: 149-172

O'kane D.J.; Lee J., 1985: Chemical characterization of lumazine protein from photobacterium leognathi comparison with lumazine protein from photobacterium phosphoreum. The properties of lumazine proteins purified from the marine bioluminescent bacteria P. phosphoreum, a psychrophile, and P. leiognathi, a relatively thermophilic species, are compared. An accurate 1:1 stoichiometry of binding of the ligand 6,7-dim...

O'Kane, D.J.; Lee, J., 1985: Chemical characterization of lumazine protein from Photobacterium leiognathi: comparison with lumazine protein from Photobacterium phosphoreum. The properties of lumazine proteins purified from the marine bioluminescent bacteria Photobacterium phosphoreum, a psychrophile, and Photobacterium leiognathi, a relatively thermophilic species, are compared. An accurate 1:1 stoichiometry of bindi...

O'Kane, D.J.; Lee, J., 1985: Physical characterization of lumazine proteins from Photobacterium. The physicochemical properties of Photobacterium lumazine proteins have been investigated. The molecular weights obtained by several physical techniques are in good agreement, and the averages are 2% and 8% higher than the minimum molecular weight...

Malak H.; Lakowicz J.R.; Lee J., 1991: Distribution of tryptophan lumazine distances in photobacterium phosphoreum lumazine protein measured by frequency domain fluorescence energy transfer. Biophysical Journal 59(2 PART 2): 477A

Illarionov, B.; Illarionova, V.; Lee, J.; van Dongen, W.; Vervoort, J., 1994: Expression and properties of the recombinant lumazine (riboflavin) protein from Photobacterium leiognathi. Photobacterium leiognathi lumazine protein has been expressed in Escherichia coli in high yield, 30 mg/l. The cloned gene was one previously reported by Illarionov (EMBL X56534), that had a similar sequence and was located in the same position as...

Illarionov, B.; Illarionova, V.; Lee, J.; Van-Dongen, W.; Vervoort, J., 1994: Expression and properties of the recombinant lumazine (riboflavin) protein from Photobacterium leiognathi. Photobacterium leiognathi lumazine protein has been expressed in Escherichia coli in high yield, 30 mg/l. The cloned gene was one previously reported by Illarionov (EMBL X56534), that had a similar sequence and was located in the same position as...

Sato, Y.; Shimizu, S.; Ohtaki, A.; Noguchi, K.; Miyatake, H.; Dohmae, N.; Sasaki, S.; Odaka, M.; Yohda, M., 2010: Crystal structures of the lumazine protein from Photobacterium kishitanii in complexes with the authentic chromophore, 6,7-dimethyl- 8-(1'-D-ribityl) lumazine, and its analogues, riboflavin and flavin mononucleotide, at high resolution. Lumazine protein (LumP) is a fluorescent accessory protein having 6,7-dimethyl-8-(1'-d-ribityl) lumazine (DMRL) as its authentic chromophore. It modulates the emission of bacterial luciferase to shorter wavelengths with increasing luminous st...