• Microbiology and Biotechnology Letters
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• v.23 no.2
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• pp.187-196
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• 1995

In order to produce fuctional chitooligosaccharides, a strain excreting mainly endo-type chitinase suitable for chitooligosaccharides production was newly screened and identified as Aspergillus fumigatus JC-19. The chitinase excretion was repressed in nutrient rich medium but stimulated by colloidal chitin indicating that the chitinase is inducible type enzyme. Maximum secretion of the enzyme was observed at pH 7.0 and 37$\circ$C . The growth and chitinase production patterns of Aspergillus fumigatus JC-19 showed that the cell growth reached maximum after 4-5 days with final chitinase concentration of 0.46 unit per ml. Excreted chitinase was purified by ammonium sulfate precipitation, colloidal chitin adsorption, anion exchange chromatography, and gel filtration, respectively, and measured M.W of 50 KDa. The enzyme reaction carried out both by crude and purified chitinase showed that the purified chitinase accumulated more chitooligosaccharides of 1-6 degree of polymerization than that of crude chitinase.

• Korean journal of food and cookery science
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• v.15 no.2
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• pp.102-107
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• 1999

Several ingredients of kimchi including chinese cabbage, garlic, leek, big green onion, and small green onion were assayed for their chitinase activities. Kimchi with various leek contents (4, 8, 12%) were fermented at 15$^{\circ}C$ for 9 days and the chitinase (EC 3.2.1.14) activity and textural properties were determined. The chitinase activity of the ingredients was in the order of garlic>leek>small green onion>chinese cabbage>big green onion. During fermentation, the chitinase activity of kimchi juice appeared more prominent than that of kimchr tissue, however, it was decreased in all kimchi samples among which the control sample showed a remarkable drop. The activity of chitinase in kimchi tissue increased until 3rd or 5th day of fermentation and then decreased. The puncture force of all kimchi samples decreased and those of leek-added kimchi were higher than those of control. The above results suggested that the addition of leek for kimchi preparation could contribute to the improvement of textural qualities of kimchi due to chitinase activities of leek during fermentation.

• KSBB Journal
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• v.16 no.4
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• pp.365-369
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• 2001

Chitinase producing microorganism, Serratia marcescens KY, was isolated from seashore mud around Beobseongpo in Chunnam province by selective enrichment culture. As the colloidal chitin concentration increased, chitinase production was increased. But chitinase production with addition of other carbon sources (glucose, fructose, galactose, maltose, sucrose, starch) was decreased. The effect of nitrogen sources on the chitinase production with serratia marcescens KY was as fellows. The opitimum mineral concentration for chitinase production was K$_2$HPO$_4$ 0.2 g/L and MgSO$_4$ 0.20 ∼ 0.25 g/L, respectively. The effect of nitrogen sources on chitinase production by Serratia marcescens KY was increased as follows, tryptone > yeast extract > beef extract > asparagine.

• Korean Journal Plant Pathology
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• v.10 no.4
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• pp.277-283
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• 1994

An antimicrobial chitinase was purified from grapefruit extract and its properties were characterized. The chitinase was purified with a single step chromatography on regenerated chitin affinity gel column. The molecular weight of the purified chitinase was 29 kDa. The grapefruit extract contained the chitinase protein more than 50% of its total soluble proteins measured by coomassie stained protein bands. When the purified chitinase was incubated with polymers of N-acetylglucosamine (NAG), such as mycelia of Fusarium oxysproum and swollen chitin, they were degraded to oligosaccharides, and the oligosaccharides were then further hydrolyzed by the same enzyme to monomer and dimer of NAG. This result suggests that the chitinase contained both endo- and exo- chitinase activities. The chitinase was stable to heat and pH treatment; its activity was not diminished by the heat treatment upto 7$0^{\circ}C$ for 1 hr, and it showed a pH stability in the range of pH 4.0 to 12.0.

• Applied Biological Chemistry
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• v.36 no.1
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• pp.1-6
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• 1993

Chitinase was induced in rice cell suspension culture with treatment of chitooligosaccharides mixture. Among eleven isozymes found in 10% polysacrylamide gel electropherogram, four isozymes were identified as induced enzymes. Acidic chitinase fraction separated in DEAE-cellulose column chromatography, includes three induced chitinase, while basic fraction contains only one induced isozyme. Treatment of chitooligosaccharides mixture enhanced the contents in both protein and chitinase activity in cell suspension culture media, but increase in chitinase activity was much higher than in protein.

• Biotechnology and Bioprocess Engineering:BBE
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• v.7 no.1
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• pp.38-42
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• 2002

A chitinase gene (pCHi58) encoding a 58 kDa chitinase was isolated from the Serratia marcescens KCTC 2172 cosmid library. The chitinase gene consisted of a 1686 bp open reading frame that encoded 562 amino acids. Escherichia coil harboring the pChi58 gene secreted a 58 kDa chitinase into the culture supernatant. The 58 kDa chitinase was purified using a chitin affinity column and mono-S column. A nucleotide and N-terminal amino acid sequence analysis showed that the 58 kDa chitinase had a leader peptide consisting of 23 amino acids which was cleaved prior to the 24th alanine. The 58 KDa chitinase exhibited a $98\%$ similarity to that of S. marcescens OMB 1466 in its nuclotide sequence. The chitinolytic patterns of the 58 kDa chitinase released N,N'-diacetyl chitobiose (NAG2) as the major hydrolysis end-product with a trace amount of N-acetylglucosamine. When a 4-methylumbellyferyl-N-acetylglucosamin monomer, dimmer, and tetramer were used as substrates, the 58 kDa chitinase did not digest the 4-Mu-NAG monomer $(analogue\;of\;NAG_2)$, thereby indicating that the 58 kDa chitinase was likely an endochitinase. The optimum reaction temperature and pH of the enzyme were $50^{\circ}C$ and 5.0, respectively.

• Journal of the Korean Chemical Society
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• v.40 no.1
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• pp.72-80
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• 1996

A chitinase-producing bacterium, Serratia marcescens JM, was isolated from a seashore muds. A chitinase was purified by ammonium sulfate precipitation, affinity adsorption, hydroxylapatite and sephadex G-200 column chromatography. The chitinase obtained from Serratia marcescens JM was purified 42.2 folds with the overall yield of 7.1%. The purified chitinase showed a single band on sodium dodecyl sulfate polyacrylamide gel electrophoresis. The molecular weight of the enzyme was 59,000 and the apparent kinetic parameters $K_m\;and\;V_{max}$ for the purified chitinase were 5.17 mg/mL and 39.8 unit/mL, respectively. The optimum pH and temperature of the purified chitinase were 7.0 and 50$^{\circ}C$, respectively and the purified enzyme was stable on pH 7.0 up to 50$^{\circ}C$. The enzyme were activated by $Cu^{2+},\;Ca^{2+}\;and\;Mg^{2+}$ and inhibited by $Hg^{2+}$ respectively. In addition, Cysteine increased the chitinase activity and EDTA, MIA, PCMB and SDS inhibited enzyme activities. Major cations, $MG^{2+},\;Ca^{2+},\;K^+\;and\; Na^+$ present in seawater slightly stimulated the chitinase activity.

• Applied Biological Chemistry
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• v.42 no.3
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• pp.181-187
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• 1999

For the selection of an effective antagonistic biocontrol agent, we have isolated an antagonistic bacterium which produced extracellular chitinase, from a local soil of Kyongju, Korea. The selected strain was identified as Serratia proteamaculans 3095. The chitinase produced from Serratia sp. 3095 showed antifungal activity which can attack the hypha surface of Fusarium oxysporum and F. solani. The carbon and nitrogen sources for chitinase production were 0.15% colloidal chitin and 0.1% ammonium sulfate, respectively. Glucose in the chitinase production medium might inhibit the production of chitinase by feed back repression. The antagonistic Serratia sp. 3095 also showed a powerful biocontrol activity against F. oxysporum through in vitro test and in vivo pot test.

• Microbiology and Biotechnology Letters
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• v.9 no.4
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• pp.179-183
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• 1981

An extracellular chitinase was purified from the culture fluid of Streptomyces sp. 115-5, and its inhibition mechanism by end product was studied. The activity of chitinase was suppressed by the reducing sugar as the reaction proceeded, and the activity was inhibited by the addition of D-glucose. Besides D-glucose, the rate of chitin hydrolysis was inhibited by D-glucuronic acid, D-sorbitol and D-xylose in the reaction system of the enzyme. it was found that the hydroxyl groups at the C-2, C-3 and C-4 position of D-glucose molecule play an important role in the inhibition of the chitinase activity. D-glucose was found to inhibit the enzyme activity by mixed type of competitive and non-competitive mode.

• Applied Biological Chemistry
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• v.41 no.2
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• pp.125-129
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• 1998

Xanthomonas sp. isolated from soil exhibited cell wall lytic activity of Candida albicans and secreted chitinase in chitin media. Especially, the chitinase activity was induced by chitin and reached a maximum level at 3 days culture in chitin media. We constructed genomic library of Xanthomonas sp. using cosmid vector in E. coli. Oligonucleotide probe was synthesized from the consensus sequence corresponding to chitinase active site, which was derived from the comparison of amino acid sequences of bacterial chitinase genes. Using this oligonucleotide probe, we screened the genomic library. By restriction enzyme mapping of the positive clones, we identified 4 independent clones which may contain the chitinase gene. One of the clones, named pXCH1 (1.2 kb insert), was further analyzed. Northern blot analysis indicated that is transcripts, 1 kb and 0.8 kb, were induced by chitin. When the cloned gene was induced by IPTG in E.coli cell, chitinase activity which was secreted onto culture media was not observed. However, when the cell was disrupted by using sonicator and then centrifuged, the supernatant exhibited chitinase activity. SDS-PAGE of the supernatant indicated that about 35 kDa protein was induced by IPTG. From these results, it was concluded that the cloned DNA was one of the chitinase genes of Xanthomonas sp.