• Proceedings of the Microbiological Society of Korea Conference
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• 2002.10a
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• pp.217.2-217.2
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• 2002

• Journal of Plant Biotechnology
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• v.35 no.1
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• pp.41-45
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• 2008

Laccase (EC 1.10.3.2) is a small group of enzymes that catalyze the oxidation of a broad range of phenolic compounds including hazardous and recalcitrant pollutants in the environment. This study attempted to develop an efficient system for production of a recombinant laccase by chloroplast genetic transformation of tobacco. Chloroplast transformation vector was constructed and introduced into the tobacco chloroplast genome using particle bombardment. Chloroplast-transformed plants were subsequently regenerated. PCR and southern blot analyses confirmed stable integration of the laccase gene into the chloroplast genome. Northern blot analysis revealed that mRNA of the laccase gene was highly expressed in chloroplast-transformed plants.

• Korean Journal of Microbiology
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• v.44 no.3
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• pp.199-202
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• 2008

A white rot fungus Irpex lacteus produced lignin degrading enzymes, which showed degrading activity against various recalcitrant compounds. However, laccase, one of the lignin degrading enzymes, was too low to be assayed by spectrophotometry using o-tolidine as the chromogenic substrate in this fungus under various culture conditions. A laccase expression vector was constructed using a cDNA from Phlebia tremellosa with the constitutively expressed promoter of glyceraldehydes-3-phosphate dehydrogenase gene, and introduced into I. lacteus by the restriction enzyme mediated integration transformation through the protoplast-$CaCl_2$ procedure. Two transformants showed highly increased laccase activities at the early growth phase in the minimal liquid medium, and they not only degraded bisphenol A, a notorious endocrine disrupting chemical, but also removed the estrogenic activity effectively.

• Journal of Microbiology and Biotechnology
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• v.29 no.9
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• pp.1383-1390
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• 2019

In this study, we expressed cotA laccase from Bacillus subtilis on the surface of B. subtilis spores for efficient decolorization of synthetic dyes. The cotE, cotG, and cotY genes were used as anchoring motifs for efficient spore surface display of cotA laccase. Moreover, a $His_6$ tag was inserted at the C-terminal end of cotA for the immunological detection of the expressed fusion protein. Appropriate expression of the CotE-CotA (74 kDa), CotG-CotA (76 kDa), and CotY-CotA (73 kDa) fusion proteins was confirmed by western blot. We verified the surface expression of each fusion protein on B. subtilis spore by flow cytometry. The decoloration rates of Acid Green 25 (anthraquinone dye) for the recombinant DB104 (pSDJH-EA), DB104 (pSDJH-GA), DB104 (pSDJH-YA), and the control DB104 spores were 48.75%, 16.12%, 21.10%, and 9.96%, respectively. DB104 (pSDJH-EA) showed the highest decolorization of Acid Green 25 and was subsequently tested on other synthetic dyes with different structures. The decolorization rates of the DB104 (pSDJH-EA) spore for Acid Red 18 (azo dye) and indigo carmine (indigo dye) were 18.58% and 43.20%, respectively. The optimum temperature for the decolorization of Acid Green 25 by the DB104 (pSDJH-EA) spore was found to be $50^{\circ}C$. Upon treatment with known laccase inhibitors, including EDTA, SDS, and $NaN_3$, the decolorization rate of Acid Green 25 by the DB104 (pSDJH-EA) spore decreased by 23%, 80%, and 36%, respectively.

• 한국균학회소식:학술대회논문집
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• 2009.05a
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• pp.51-52
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• 2009

• Journal of Microbiology and Biotechnology
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• v.24 no.4
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• pp.545-555
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• 2014

It has been previously demonstrated that laccases exhibit great potential for use in several industrial and environmental applications. In this paper, two laccase isoenzyme genes, lccB and lccC, were cloned and expressed in Pichia pastoris GS115. The sequence analysis indicated that the lccB and lccC genes consisted of 1,563 and 1,584 bp, and their open reading frames encoded 520 and 527 amino acids, respectively. They had 72.7% degree of identity in nucleotides and 86.7% in amino acids. The expression levels of LccB and LccC were up to 32,479 and 34,231 U/l, respectively. The recombinant laccases were purified by ultrafiltration and $(NH_4)_2SO_4$ precipitation, showing a single band on SDS-PAGE, which had a molecular mass of 58 kDa. The optimal pH and temperature for LccB were 2.0 and $55^{\circ}C$ with 2,2'-azinobis-[ 3-ethylbenzthiazolinesulfonic acid (ABTS) as a substrate, whereas LccC exhibited optimal pH and temperature at 3.0 and $60^{\circ}C$. The apparent kinetic parameters of LccB were 0.43 mM for ABTS with a $V_{max}$ value of 51.28 U/mg, and the Km and $V_{max}$ values for LccC were 0.29 mM and 62.89 U/mg. The recombinant laccases were able to decolorize five types of dyes. Acid Violet 43 (100 g/ml) was completely decolorized by LccB or LccC (2 U/ml), and the decolorization of Reactive Blue KN-R (100 g/ml) was 91.6% by LccC (2 U/ml). Thus, the study characterizes useful laccase isoenzymes from T. versicolor that have the capability of being incorporated into the treatment of similar azo and anthraquinone dyes from dyeing industries.

• Proceedings of the Microbiological Society of Korea Conference
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• 2005.05a
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• pp.217.3-217
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• 2005

• Proceedings of the Zoological Society Korea Conference
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• 1997.10b
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• pp.243.1-243
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• 1997

No Abstract, See Full Text

• Proceedings of the Microbiological Society of Korea Conference
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• 2005.05a
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• pp.159-161
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• 2005

Hypovirus infection of the chestnut blight fungus Cryphonectria parasitica is a useful model system to study the hypoviral regulation of fungal gene expression. The hypovirus is known to downregulate the fungal laccase1 (lac 1), the modulation of which is tightly governed by the inositol triphosphate ($IP_3$) and calcium second messenger system in a virus-free strain. We cloned the gene cplc1 encoding a phosphatidyl inositol-specific phospholipase C (PLC), in order to better characterize the fungal gene regulation by hypovirus. Sequence analysis of the cplc1 gene indicated that the protein product contained both the X and Y domains, which are the two conserved regions found in all known PLCs, with a 133 amino acid extension between the 2nd ${\beta}$-strand and the ${\alpha}$-helix in the X domain. In addition, the gene organization appeared to be highly similar to that of a ${\delta}$ type PLC. Disruption of the cplc1 gene resulted in slow growth and produced colonies characterized by little aerial mycelia and deep orange in color. In addition, down regulation of lac1 expression was observed. However, temperature sensitivity, osmosensitivity, virulence, and other hypovirulence-associated characteristics did not differ from the wild-type strain. Functional complementation of the cplc1-null mutant with the PLC1 gene from Saccharomyces cerevisiae restored lac1 expression, which suggests that the cloned gene encodes PLC activity. The present study indicates that the cplc1 gene is required for appropriate mycelial growth, and that it regulates the lac1 expression, which is also modulated by the hypovirus. Although several PLC genes have been identified in various simple eukaryotic organisms, the deletion analysis of the cplc1 gene in this study appears to be the first report on the functional analysis of PLC in filamentous fungi.

• Molecules and Cells
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• v.25 no.1
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• pp.112-118
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• 2008

Laccases are multicopper-containing oxidases that catalyze the oxidation of many aromatic compounds with concomitant reduction of oxygen to water. Interest in this enzyme has arisen in many fields of industry, including detoxification, wine stabilization, paper processing, and enzymatic conversion of chemical intermediates. In this study, we cloned a laccase gene (GLlac1) from the white-rot fungus Ganoderma lucidum. The cloned gene consists of 4,357 bp, with its coding region interrupted by nine introns, and the upstream region has putative CAAT and TATA boxes as well as several metal responsive elements (MREs). We also cloned a full-length cDNA of GLlac1, which contains an uninterrupted open reading frame (ORF) of 1,560 bp coding for 520 amino acids with a putative 21-residue signal sequence. The DNA and deduced amino acid sequences of GLlac1 were similar but not identical to those of other fungal laccases. GLlac1 was released from the cells when expressed in P. pastoris, and had high laccase activity. In addition, GLlac1 conferred antioxidative protection from protein degradation, and thus may be useful in bio-medical applications.