• BMB Reports
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• 제39권5호
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• pp.578-585
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• 2006

The cDNA of a firefly luciferase from lantern mRNA of Lampyroidea maculata has been cloned, sequenced and functionally expressed. The cDNA has an open reading frame of 1647 bp and codes for a 548-residue-long polypeptide. Noteworthy, sequence comparison as well as homology modeling showed the highest degree of similarity with H. unmunsana and L. mingrelica luciferases, suggesting a close phylogenetic relationship despite the geographical distance separation. The deduced amino acid sequence of the luciferase gene of firefly L. maculata showed 93% identity to H. unmunsana. Superposition of the three-dimensional model of L. maculata luciferase (generated by homology modeling) and three dimensional structure of Photinus pyralis luciferase revealed that the spatial arrangements of Luciferin and ATP-binding residues are very similar. Putative signature of AMP-binding domain among the various firefly species and Lampyroidea maculata was compared and a striking similarity was found. Different motifs and sites have been identified in Lampyroidea maculata by sequence analysis. Expression and purification of luciferase from Lampyroidea maculata was carried out using Ni-NTA Sepharose. Bioluminescence emission spectrum was similar to Photinus pyralis luciferase.

• International Journal of Industrial Entomology and Biomaterials
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• 제1권1호
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• pp.53-58
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• 2000

A cDNA encoding the luciferase of firefly Luciola lateralis was cloned downstream from the polyhedrin gene promoter of Bombyx mori nucleopolyhedrovirus and expressed in B. mori cells (BmN-4). The coding soquence for luciferase was inserted into pBmKSK2 rectors) which was reconstructed from the polyhedrin-based transfer vector pBmKSKl by modifying cloning sites. Recombinant virus, BmK2-LUCDF, containing the luciferase gene was selected and purified in BmN-4 cells. The emission of luminescence by luciferase was only detected in BmK2-LUCDF-infected cell extracts. This result indicates that the cloned new luciferase gene of firefly L. lateralis can be expressed efficiently in baculovirus expression system and used as a useful reporter gene.

• International Journal of Industrial Entomology and Biomaterials
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• 제7권2호
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• pp.155-159
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• 2003

We describe here the complete nucleotide sequence and the exon-intron structure of the luciferase gene of the firefly, Lampyris noctiluca. The luciferase gene of the L. noctiluca firefly consisted of six introns and seven exons coding for 547 amino acid residues. From the translational start site to the end of last exon, the genomic DNA length of the L. noctiluca luciferase gene spans 1,976 bp.

• Asian-Australasian Journal of Animal Sciences
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• 제11권5호
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• pp.498-502
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• 1998

The present study was designed, fIrst to develop the new methodology to measure the bioluminescence activity easily in live developing bovine embryos by photoncounting, and secondly to compare the expression efficiency of four luciferase reporter genes in bovine embryos at four- to 16-cell stages. In experiment 1, equimolar pSVlacZ and pSVEluc were microinjected into the pronucleus of fertilized bovine oocytes. At 2 days after micro injection, bioluminescence activity of these embryos was measured by photoncounting with a luminometer for 1 min, and lacZ gene expression in the same embryos was assayed by X-gal staining. All the luciferase-positive oocytes showed some bacterial ${\beta}$-galactosidase activity irrespective of the intensity. In experiment 2, four firefly luciferase genes (pTKEluc, pTK6WEluc, pSVEluc and pMiwluc) were introduced by micro injection, and the injected embryos were cultured for the following 2 days. Detection of the luciferase gene expression was done by photoncounting at 5 to 55 min. Over the measurement period, the luciferase activity was almost constant irrespective of the transgenes microinjected. The luciferase activity and expression efficiency at 2 days after microinjection were not significantly affected by the difference in the microinjected transgenes. The present results demonstrated that the bioluminescence activity in live developing bovine embryos could be measured quickly by photoncounting.

• International Journal of Industrial Entomology and Biomaterials
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• 제9권2호
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• pp.155-165
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• 2004

In Korean Peninsula including neighboring islands and Japanese Islands identical firefly species or the species belonging to same genera occur together in both territories. These geographic firefly species, nonetheless, have never been subject to taxonomic consideration together until recently, lacking clear species status and phylogenetic relationships. A recent serial study of these fireflies using luciferase gene and/or portions of mitochondrial DNA sequences provided some insight into these populations in terms of validity of species name, phylogenetic relationships, and speciation event. In this article, thus, we have reviewed the recent progress on phylogenetic and/or population genetic aspects of these species, i.e., Hotaria-group fireflies, Luciola lateralis, and Pyrocoelia rufa to better understand the firefly species in these regions.

• International Journal of Industrial Entomology and Biomaterials
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• 제7권2호
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• pp.181-189
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• 2003

We describe here the complete nucleotide sequence and the exon-intron structure of the luciferase gene of the firefly, Pyrocoelia rufa. The luciferase gene of the P. rufa firefly consisted of six introns and seven exons coding for 548 amino acid residues. From the translational start site to the end of last exon, however, the genomic DNA length of the P. rufa luciferase gene from the Korean and Chinese samples spans 1,968 bp and 1983 bp, respectively, and 3 amino acid residues were different to each other. Additionally, we also analyzed mitochondrial cytochrome oxidase I(COI) gene of the Chinese P. rufa fireflies. Analysis of DNA sequences from the mitochondrial COI protein-coding gene revealed 4 mitochondrial DNA sequence-based haplotypes with a maximum divergence of 0.7％. With the 20 P. rufa haplotypes found in Korea, phylogenetic analyses using PAUP and PHYLIP subdivided the P. rufa into three clades, termed clades A and B for the Korean sample, and clade C for the Chinese sample.

• International Journal of Industrial Entomology and Biomaterials
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• 제2권2호
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• pp.91-105
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• 2001

Taxonomic and phylogenetic studies of firefly in Japan have been reviewed. Fourty-six lampyrid species and one rhagophthalmid are distributed in the Japanese Islands including the Ryukyus. Recently, molecular phylogenetic approaches have been employed in the systematic study of firefly using mitochondrial and luciferase genes. Based on the molecular phylogenetic trees, evolutionary process of flashing patterns related strictly to mating behavior was estimated. Furthermore, genetic diversity studies revealed geographic differentiation patterns within species, and conservation measures of firefly were proposed to protect genetic resources endemic to the localities.

• Bulletin of the Korean Chemical Society
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• 제35권2호
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• pp.425-430
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• 2014

Amyloidogenic proteins often exhibit fibrillar polymorphism through alternative assembly processes, which has been considered to have possible pathological implications. Here, firefly luciferase (LUC) is shown to induce amyloid polymorphism of ${\alpha}$-synuclein, the major constituent of Lewy bodies found in Parkinson's disease, by acting as a novel template. The drastically accelerated fibrillation kinetics of ${\alpha}$-synuclein with LUC required the nucleation center produced by the active enzyme of LUC. Fluorescent dye binding, transmission electron microscopy, and Fourier transformed infrared spectroscopy revealed the morphologically distinctive amyloid fibrils of ${\alpha}$-synuclein prepared in the absence or presence of LUC. As the altered morphological characteristics became inherent to the mature fibrils, those properties were inherited to next-generations via nucleation-dependent fibrillation process. The seed control, therefore, would be an effective means to modify amyloid fibrils with different biochemical characteristics. In addition, the LUC-directed amyloid fibrillar polymorphism also suggests that other cellular biomolecules including enzymes in general are able to diversify amyloid fibrils, which could be self-propagated with diversified biological activities, if any, inside cells.

• BMB Reports
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• 제44권2호
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• pp.102-106
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• 2011

Thermodynamic stability and refolding kinetics of firefly luciferase and three representative mutants with depletion of negative charge on a flexible loop via substitution of Glu by Arg (ER mutant) or Lys (EK mutant) as well as insertion of another Arg in ER mutants (ERR mutant) was investigated. According to thermodynamic studies, structural stability of ERR and ER mutants are enhanced compared to WT protein, whereas, these mutants become prone to aggregation at higher temperatures. Accordingly, it was concluded that enhanced structural stability of mutants depends on more compactness of folded state, whereas aggregation at higher temperatures in mutants is due to weakening of intermolecular repulsive electrostatic interactions and increase of intermolecular hydrophobic interactions. Kinetic results indicate that early events of protein folding are accelerated in mutants.

• Nuclear Medicine and Molecular Imaging
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• 제41권1호
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• pp.54-58
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• 2007

Purpose: Luciferase is one of the most commonly used reporter enzymes in the field of in vivo optical imaging. D-luciferin, the substrate for firefly luciferase has very high cost that allows this kind of experiment limited to small animals such as mice and rats. In this current study, we validated local injection of D-luciferin in the articular capsule for bioluminescence imaging in rabbits. Materials and Methods: Chondrocytes were cultured and infected by replication-defective adenoviral vector encoding firefly luciferase (Fluc). Chondrocytes expressing Fluc were injected or implanted in the left knee joint. The rabbits underwent optical imaging studies after local injection of D-luciferin at 1, 5, 7, 9 days after cellular administration. We sought whether optimal imaging signals was could be by a cooled CCD camera after local injection of D-luciferin. Results: Imaging signal was not observed from the left knee joint after intraperitoneal injection of D-luciferin (15 mg/kg), whereas it was observed after intraarticular injection. Photon intensity from the left knee joint of rabbits was compared between cell injected and implanted groups after intraarticular injection of D-luciferin. During the period of imaging studies, photon intensity of the cell implanted group was 5-10 times higher than that of the cell injected group. Conclusion: We successfully imaged chondrocytes expressing Fluc after intraarticular injection of D-luciferin. This technique may be further applied to develop new drugs for knee joint disease.