• YAKHAK HOEJI
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• v.43 no.4
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• pp.502-508
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• 1999

Chitin is an important structural component of fungal cell wall and is synthesized by chitin synthase I, II, and III. The chitin synthase II is an essential enzyme for the formation of primary septum in Saccharomyces cerevisiae. Therefore, specific inhibitors of this enzyme might block the formation of fungal cell wall and could be used as effective antifungal agents. To search chitin synthase IIinhibitors from natural products, 67 plants were extracted with methanol and examined for the inhibitory activities against chitin synthase II of S. cerevisiae by our cell free assay system. As a result, the extracts from 16 plants showed more than 70% inhibition at the concentration of $280{\;}\mu\textrm{g}/ml$. Of note, Laurus nobilis (81.4%), Lonicera maackii (81.5%), Berchemia berchemiaefolia (82.9%), Koelreuteria paniculata (87.9%), Chamaecyparis pisifera (86%) and Taxus cuspidata (83.9%) inhibited strogly the chitin synthase IIactivity.

• Journal of Microbiology and Biotechnology
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• v.10 no.2
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• pp.251-257
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• 2000

Chitin synthases are identified as key enzymes of chitin biosynthesis in most of the fungi. Among them, chitin synthase II has been reported to be and essential enzyme in chitin biosynthesis, and exists as a membrane-bound form. To search and screen new antifungal agents from natural resources to inhibit chitin synthase II, the assay conditions were established using the enzyme isolated from Saccharomyces cerevisiae ECY38-38A(pAS6) that overproduces only chitin synthase II. This enzyme was activated only by partial proteolysis with trypsin. Its actibity reached the maximum at $80{\;}\mu\textrm{g}/ml$ of trypsin and was strongly stimulated by 2.0 mM $Co^{2+}$, 1.0 nM UDP-[$^{14}C$]-GicNAc, and 32 mM free-GlcNAc. Under these assay conditions, the highest chitin synthase II activity was observed by incubation at $30^{\circ}C$ for 90 min. However, and extremely narrow range of organic solvents up to as much as 25% of DMSO and 25% of MeOH was useful for determining optimal assay conditions. After a search or potent inhibitors of chitin synthase II from natural resources, prodigiosin was isolated from Serratia marcescens and purified by solvent extration and silica gel column chromatographies. The structure of prodigiosin was determined by UV, IR, Mass spectral, and NMR spectral analyses. Its molecular weight and formula were found to be 323 and $C_{20}H_{25}N_{3}O$, respectively. Prodigiosin ingibited chitin synthase II by 50% at the concentration of $115{\;}\mu\textrm{g}/ml$.

• YAKHAK HOEJI
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• v.43 no.4
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• pp.509-515
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• 1999

To search for new chitin biosynthesis inhibitors from natural sources, several higher plants were examined the inhibitory activity against chitin synthase IIby enzymatic assay. Among them, the extract of Schizandra chinensis strongly showed the inhibitory activity against chitin synthase II. Gomisin N and wuweizisu C were isolated from Schizandra chinensis and showed $IC_50$ value of $62.4{\;}\mu\textrm{g}/ml$ and $19.2{\;}\mu\textrm{g}/ml$, respectively. Activities of these compounds were more stronger than that of polyxin D. However, gomisin N and wuweizisu C showed weakly antifungal activities against various human pathogens.

• Journal of Microbiology
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• v.35 no.3
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• pp.159-164
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• 1997

The phylogenetic study of Penicilium chrysogenum was performed based on amino acid sequence comparison of chitin synthase. Phylogenetic trees were constructed with the deduced amino acid sequences of the highly conserved region of chitin synthease gene fragments amplified by PCR. The BlasP similarity searcch and the bootstrap analysis of the deduced amino acid sequences of chitin synthase from P. chrysogenum with those form other fungi showed a close evolutionary relationship of Penicillium to ascomycetous fungi, especially to genus Aspergilus. The result from bootstrap analysis of the deduced amino acid sequences of the Class II chitin synthase from ascomyceteous fungi supported the usefulness of the Class II chitin synthease for phylogenetic study of filamentous fungi.

• Mycobiology
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• v.29 no.4
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• pp.179-182
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• 2001

Chitin synthases(UDP-N-acetyl-D-glucosamine: chitin 4-$\beta$-N-acetyl-D-glucosaminyl transferase, EC 2.4.1.16) catalyze the synthesis of chitin from UDP-N-acetyl-D-glucosamine. Two zymogenic type of chitin synthase gene(TmCHS1 and TmCHS2) were amplified and its nucleotide sequences were determined. By the amino acid comparison and UPGMA tree grouping, TmChs1 and TmChs2 were classified as class II and class IV chitin synthases respectively. The class II type TmChs1 was grouped with others of Agaricales ectomycorrhizal mushroom. Additionally the phylogenetic tree was well adapted to Hymenomycete previously classified by morphological and physiological characteristics.

• The Korean Journal of Mycology
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• v.25 no.3 s.82
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• pp.167-175
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• 1997

The PCR fragments of two distinct chitin synthase genes, PdCHSl and PdCHS2, were cloned from Penicillium diversum KCTC 6786. The nucleotide sequences of PdCHSl and PdCHS2 contained uninterrupted open reading frames (ORFs) of 570 bp excluding the primer sequence. The similarity analysis of the deduced amino acid sequences using BLASTP indicated that the possible evolutionary relationship between P. diversum and ascomycetous fungi. Multialignment of the deduced amino acid sequences of PdCHSs using CLASTAL W revealed that the PdCHSs fell into two different classes: PdCHSl into Class I and PdCHS2 into Class II of chitin synthase defined by Bowen et al. (1992). By Southern blot analysis, it was shown that each of the two genes is present as a single copy in the genome of P. diversum KCTC 6786.

• Journal of Microbiology
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• v.35 no.3
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• pp.222-227
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• 1997

DNA fragments homologous to chitin synthase gene were amplified from the genomic DNA of Beauveria brongniartii by PCR using degenerate primers. Cloning and sequencing of the PCR-amplified fragments led to the identification of a gene, designated BbCHSl. Comparison of the deduced amino acid sequence of BbCHSl with those of other Euascomycetes revealed that BbCHSl is a gene for class II chitin synthase. The Blastp search of the deduced amino acid sequence of BbCHSl displayed the highest rate of similarity, 95.8%, with CHS2 of Metarhizium unisopliae. Phylogenetic analysis of the amino acid sequences confirmed the taxonomic and evolutionary position of B. brongniartii, which was previously derived by traditional fungal classification based on morphological features.

• Bulletin of the Korean Chemical Society
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• v.30 no.12
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• pp.3092-3094
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• 2009

• Korean Journal of Environmental Biology
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• v.23 no.4
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• pp.335-342
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• 2005

We cloned seven genes encoding chitin synthases (CHSs) by PCR amplification from genomic DNAs of four strains of the genus Sporobolomyces and of Bensingtonia subrosea using degenerated primers based on conserved regions of the CHS genes. Though amino acid sequences of these genes were shown similar as 176 to 189 amino acids except SgCHS2, DNA sequences were different in size, which was due to various introns present in seven fragments. Alignment and phylogenetic analysis of their deduced amino acid sequences together with the reported CHS genes of basidiomycetes separated the sequences into classes I, II and III. This analysis also permitted the classification of isolated CHSs; SgCHS1 belongs to class I, BsCHS1, SaCHS1, SgCHS2, SpgCHS1, and SsCHS1 belong to class II, and BsCHS2 belongs to class III. The deduced amino acid sequences involving in class II that were discovered from five strains were also compared with those of other basidiomycetes by CLUSTAL X program. The bootstrap analysis and phylogenetic tree by neighbor-joining method revealed the taxonomic and evolutionary position for four strains of the genus Sporobolomyces and for Bensingtonia subrosea which agreed with the previous classification. The results clearly showed that CHS fragments could be used as a valuable key for the molecular taxonomic and phylogenetic studies of basidiomycetes.

• Mycobiology
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• v.38 no.2
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• pp.102-107
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• 2010

We identified a gene for $\beta$-1,3-glucan synthesis (GBG1), a nonessential gene whose disruption alters cell wall synthesis enzyme activities and cell wall composition. This gene was cloned by functional complementation of defects in $\beta$-1,3-glucan synthase activity of the the previously isolated Saccharomyces cerevisiae mutant LP0353, which displays a number of cell wall defects at restrictive temperature. Disruption of the GBG1 gene did not affect cell viability or growth rate, but did cause alterations in cell wall synthesis enzyme activities: reduction of $\beta$-1,3-glucan synthase and chitin synthase III activities as well as increased chitin synthase I and II activities. GBG1 disruption also showed altered cell wall composition as well as susceptibility toward cell wall inhibitors such as Zymolyase, Calcofluor white, and Nikkomycin Z. These results indicate that GBG1 plays a role in cell wall biogenesis in S. cerevisiae.