• Journal of Microbiology
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• v.43 no.6
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• pp.487-492
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• 2005

This study demonstrated that the brown rot basidiomycete Fomitopsis palustris was able to degrade crystalline cellulose (Avicel). This fungus could also produce the three major cellulases (exoglucanases, endoglucanases, and $\beta-glucosidase$) when the cells were grown on $2.0\%$ Avicel. Avicel degraded by F. palustris showed a decrease in relative crystallinity from $83\%\;to\;78.5\%$ after 14 days of incubation. The characterization study indicated that optimum pH was 4.5 and optimum temperature was $70^{\circ}C$ for exoglucanase (cellobiohydrolase) activity. Hydrolysis of Avicel by the crude enzyme from F. palustris yielded 1.6 mg/ml of glucose after 43 h, which corresponded to a cellulose conversion degree of $3.2\%$. Therefore, this study revealed for the first time that the brown rot basidiomycete F. palustris produces cellulases capable of yielding soluble sugars from crystalline cellulose.

• Journal of Microbiology and Biotechnology
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• v.25 no.7
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• pp.1101-1107
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• 2015

The role of CRE1 in a thermophilic fungus, Myceliophthora thermophila ATCC42464, was studied using RNA interference. In the cre1-silenced strain C88, the filter paper hydrolyzing activity and β-1,4-endoglucanase activity were 3.76-, and 1.31-fold higher, respectively, than those in the parental strain when the strains were cultured in inducing medium for 6 days. The activities of β-1,4-exoglucanase and cellobiase were 2.64-, and 5.59-fold higher, respectively, than those in the parental strain when the strains were cultured for 5 days. Quantitative reverse-transcription polymerase chain reaction showed that the gene expression of egl3, cbh1, and cbh2 was significantly increased in transformant C88 compared with the wild-type strain. Therefore, our findings suggest the feasibility of improving cellulase production by modifying the regulator expression, and an attractive approach to increasing the total cellulase productivity in thermophilic fungi.

• Journal of Microbiology and Biotechnology
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• v.17 no.11
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• pp.1811-1817
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• 2007

Extracellular enzymes from Lentinus edodes M290 on normal woods (Quercus mongolica) and waste logs from oak mushroom production were comparatively investigated. Endoglucanase, cellobiohydrolase, ${\beta}$-glucosidase, and xylanase activities were higher on waste mushroom logs than on normal woods after 1. edodes M290 inoculation. Xylanase activity was especially different, with a three times higher activity on waste mushroom logs. When the waste mushroom logs were used as a carbon source, a new 35 kDa protein appeared. After the purification, the optimal pH and temperature for xylanase activity were determined to be 4.0 and $50^{\circ}C$, respectively. More than 50% of the optimal xylanase activity was retained when the temperature was increased from 20 to $60^{\circ}C$, after a 240 min reaction. At $40^{\circ}C$, the xylanase maintained 93% of the optimal activity, after a 240 min reaction. The purified xylanase showed a very high homology to the xylanase family 10 from Aspergillus terreus by LC/MS-MS analysis. The highest Xcorr (1.737) was obtained from the peptide KWI SQGIPIDGIG SQTHLGSGGS WTVK originated from Aspergillus terreus, indicating that the 35 kDa protein was xylanase. This protein showed low homology to a previously reported L. edodes xylanase sequence.

• Korean Journal of Microbiology
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• v.51 no.3
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• pp.209-220
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• 2015

In this study, gut bacterial communities in xylophagous insects were analyzed using the pyrosequencing of 16S rRNA genes for their potential biotechnological applications in lignocelluloses degradation. The result showed that operational taxonomic units (OTUs), species richness and diversity index were higher in the hindgut than in the midgut of all insect samples analyzed. The dominant phyla or classes were Firmicutes (54.0%), Bacteroidetes (14.5%), ${\gamma}-Proteobacteria$ (12.3%) in all xylophagous insects except for Rhinotermitidae. The principal coordinates analysis (PCoA) showed that the bacterial community structure mostly clustered according to phylogeny of hosts rather than their habitats. In our study, the two carboxymethyl cellulose (CMC)-degrading isolates which showed the highest enzyme activity were most closely related to Bacillus toyonensis $BCT-7112^T$ and Lactococcus lactis subsp. hordniae $NCDO\;2181^T$, respectively. Cellulolytic enzyme activity analysis showed that ${\beta}-1,4-glucosidase$, ${\beta}-1,4-endoglucanase$ and ${\beta}-1,4-xylanase$ were higher in the hindgut of Cerambycidae. The results demonstrate that xylophagous insect guts harbor diverse gut bacteria, including valuable cellulolytic bacteria, which could be used for various biotechnological applications.

• Journal of Microbiology and Biotechnology
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• v.23 no.3
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• pp.351-356
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• 2013

A high ${\beta}$-glucosidase (BGL)-producing strain, Stereum hirsutum, was identified and isolated and showed a maximum BGL activity (10.4 U/ml) when cultured with Avicel and tryptone as the carbon and nitrogen sources, respectively. In comparison with other BGLs, BGL obtained from S. hirsutum showed a higher level of activity to cellobiose ($V_{max}$ = 172 U/mg, and $k_{cat}$ = 281/s). Under the optimum conditions (600 rpm, $30^{\circ}C$, and pH 6.0), the maximum BGL activity of 10.4 U/ml with the overall productivity of 74.5 U/l/h was observed. BGL production was scaled up from a laboratory scale (7-L fermenter) to a pilot scale (70-L fermenter). When S. hirsutum was cultured in fed-batch culture with rice straw as the carbon source in a 70-L fermenter, a comparable productivity of 78.6 U/l/h was obtained. Furthermore, S. hirsutum showed high levels of activity of other lignocellulases (cellobiohydrolase, endoglucanase, xylanase, and laccase) that are involved in the saccharification of biomasses. Application of S. hirsutum lignocellulases in the hydrolysis of Pinus densiflora and Catalpa ovata showed saccharification yields of 49.7% and 43.0%, respectively, which were higher than the yield obtained using commercial enzymes.

• Journal of Microbiology and Biotechnology
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• v.17 no.8
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• pp.1291-1299
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• 2007

Two ${\beta}$-1,4-glucanases (DI and DIII fractions) were purified to homogeneity from the culture filtrate of a cellulolytic bacteria, Cellulomonas sp. CS 1-1, which was classified as a novel species belonging to Cellulomonas uda based on chemotaxanomic and phylogenetic analyses. The molecular mass was estimated as 50,000 Da and 52,000 Da for DI and DIII, respectively. Moreover, DIII was identified as a glycoprotein with a pI of 3.8, and DI was identified as a non-glycoprotein with a pI of 5.3. When comparing the ratio of the CMC-saccharifying activity and CMC-liquefying activity, DI exhibited a steep slope, characteristic of an endoglucanase, whereas DIII exhibited a low slope, characteristic of an exoglucanase. The substrate specificity of the purified enzymes revealed that DI efficiently hydrolyzed CMC as well as xylan, whereas DIII exhibited a high activity on microcrystalline celluloses, such as Sigmacells. A comparison of the hydrolysis patterns for pNP-glucosides (DP 2-5) using an HPLC analysis demonstrated that the halosidic bond 3 from the nonreducing end was the preferential cleavage site for DI, whereas bond 2, from which the cellobiose unit is split off, was the preferential cleavage site for DIII. The partial N-terminal amino acid sequences for the purified enzymes were $^1Ala-Gly-Ser-Thr-Leu-Gln-Ala-Ala-Ala-Ser-Glu-Ser-Gly-Arg-Tyr^{15}$-for DI and $^1Ala-Asp-Ser-Asp-Phe-Asn-Leu-Tyr-Val-Ala-Glu-Asn-Ala-Met-Lys^{15}$-for DIII. The apparent sequences exhibited high sequence similarities with other bacterial ${\beta}$-1,4-glucanases as well as ${\beta}$-1,4-xylanases.

• 보존과학연구
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• s.19
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• pp.3-22
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• 1998

The effect of pH on degradation of paper by some fungi, which able to degrade cellulose, was investigated. Trichoderma koningii, Aspergillus nigerand Penicillium nigulosum were cultured at $28^{\circ}C$ for 16 days in the selective medium (PH3, PH4, PH5, PH6, PH7, PH8, PH9, PH10, PHC) containing paper as substrate. Each paper was pretreated with each pH buffer (pH 3∼pH 10, D.W.)prior to addition to the selective medium. Enzyme activities in the each culture medium were measured spectroph to metrically using C.M.C., Avicel, PNPG as the substrates for endoglucanase, exoglucanase and $\beta$-glucosidase, respectively. In all experimental fungi, the enzyme activities of PH3 and PH9 medium were usually much higher than those of other experimental groups. However in the PH6medium, enzyme activity was lower than other groups. To analyze the concentration and pattern of protein in the each culture medium, the medium was concentrated by lyophilization. The protein concentration of PH3 and PH9 medium were relatively high (T.koningii; 6.31mg, 6,19mg, A.niger; 1.62mg, 1.96mg, P.nigulosum;2.50mg, 2.73mg, respectively), but that of PH6 was relatively low. The protein pattern of each medium was analyzed by using SDS-PAGE and VDS Image Master Analysis Program. The concentrations of bands in the each lane were usually high at lane2 (PH3) and lane8 (PH9) and low at lane5 (PH6). Therefore, the incresed cellulolytic activity of fungus against acidified paper could be result of structural change and deterioration of paper caused by being acidified.

• Microbiology and Biotechnology Letters
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• v.20 no.5
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• pp.505-510
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• 1992

An endo-$\beta$-1,4-glucanase gene of Clostridium thermocellum was cloned in Escherichia coli and was considered as a novel gene by comparison with the restriction patterns of the C. thermocellum cellulase genes so far reported. The endoglucanase from recombinant E. coli was purified by column chromatography after heat treatment. The purified enzyme was a monomer having molecular weight of 40,000. The enzyme hydrolyzed CMC to glucose and cello-oligosaccharides at :naximum activities at pH 5.0 and $65^{\circ}C$. One of the endproducts, glucose, showed no inhibitory effect on the enzyme activity, while the other endproduct, cellobiose, inhibited slightly. The values of $K_{m}$ and $V_{max}$ of the enzyme for CMC were 0.39% (w/v) and 268 Ulmg protein, respectively.

• Proceedings of the Korean Society of Sericultural Science Conference
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• 2003.04a
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• pp.77-78
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• 2003

A novel -1, 4-endoglucanase (EGase, EC 3.2.1.4) cDNA belonging to glycoside hydrolase family (GHF) 45 was cloned from the mulberry longicorn beetle, Apriona germari. The cDNA encoding EGase of A. germari (Ag-EGase) is 711 base pairs long with an open reading frame of 237 amino acid residues. The deduced protein sequence of Ag-EGase showed 54% and 48% identity to phytophagous beetle Phaedon cochleariae and termite Reticulitermes speratus hindgut symbiont, respectively. (omitted)

• Proceedings of the Korean Society for Applied Microbiology Conference
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• 2000.04a
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• pp.78-89
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• 2000

Protein secretion in filamentous fungi has been shown to be restricted to actively growing hyphal tips. To determine whether an increase in the amount of growing surface area of a fungus can lead to an increase in the amount of protein secretion, we examined secretion in a temperature-sensitive Neurospora crassa mcb mutant that shows a loss of growth polarity when incubated at restrictive-temperature. Incubation of the mcb mutant at restrictive-temperature results in a three- to five-fold increase in the level of extracellular protein and a 20- fold increase in carboxymethyl cellulase activity relative to a wild-type strain. A mutation in the cr-l gene has been shown previously to suppress the apolar growth phenotype of the mcb mutant, and we find that the level of extracellular protein produced by a mcb; cr-l double mutant was reduced to that of the wild-type control. Immunolocalization of a secreted endoglucanase revealed that proteins are secreted mainly at hyphal tips in hyphae exhibiting polar growth and over the entire surface area of bulbous regions of hyphae that are produced following a shift of the mcb mutant to restrictive-temperature. These results support the hypothesis that secretion of extracellular protein by a filamentous fungus can be significantly increased by mutations that alter growth polarity.