• 한국식물병리학회지
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• 제11권3호
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• pp.258-264
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• 1995

Production and some properties of chitinolytic enzymes were investigated by 80% ammonium sulfate precipitates (crude enzymes) from culture supernatant of antagonistic bacteria, Chromobacterium violaceum strain C-61 and strain C-72, Aeromonas hydrophila, Aeromonas caviae, and Serratia marcescens. The maximum production of chitinase was obtained from the 3-day culture at 28$^{\circ}C$ in C. violaceum stains, the 6-day culture in S. marcescens, and the 2-day culture in A. hydrophila and A. caviae. In the optimum culture periods, chitinase activity of C. violaceum strains C-61 was 1.5, 5.5, 12.0 and 11.3 times higher than those of strain C-72, S. marcescens, A. hydrophila and A. caviae, respectively. However, N,N'-diacetylchitobiase activity was 3.2 times higher in S. marcescens than in C. violaceum strain C-61, and that of Aeromonas spp.was very low. On gels containing glycol chitin, chitinase of C. violaceum strains showed four isoforms of 54-, 52-, 50- and 37-kDa, whereas there were four isoforms of 58-, 52-, 48- and 38-kDa in S. arcescens, three isoforms of 70-, 58- and 54-kDa in A. hydrophila and six isoforms of 90-, 79-, 71-, 63-, 58- and 38-kDa in A. caviae. The chitinase of C. violaceum strain C-61 was most active at pH 7.0 and at 5$0^{\circ}C$ and was stable in ranges of pH 5.0~10.0 for 2 hours and of 0~5$0^{\circ}C$ for 30 min.

• 미생물학회지
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• 제44권3호
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• pp.258-263
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• 2008

대표적인 식물병원성 곰팡이 인 Phytophthora infestans와 Fusarium oxysporum의 생장을 저해하는 근권세균들을 토양에서 분리하여 동정하였으며 이 균주들이 분비하는 항진균성 물질인 siderophore, $\beta-1$,3-glucanase, hydrogen cyanide와 chitinase의 생성능을 조사하였다. 분리균주 중 Bacilus sp. RFO41은 F. oxysporum의 생장을 가장 효율적으로 억제하였으며, siderophore 생성능과 $\beta-1$,3-glucanase의 활성이 가장 우수하였다. 또 다른 분리균주인 Bacilus sp. PS2는 P. infestans의 생장을 가장 많이 억제하였으며, chitinase 활성과 hydrogen cyanide 생성능이 가장 우수하였다. F. oxysporum과 P. infestans에 대한 항진균 효과는 근권세균이 생산하는 siderophore, $\beta-1$,3-glucanase, hydrogen cyanide와 chitinase의 활성에 따라 차이가 있음을 알 수 있었다.

• 한국미생물·생명공학회지
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• 제21권1호
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• pp.6-12
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• 1993

토양에서 분리한 294균주 중에서 chitinase 생산성이 우수한 균주를 선택하였으며 형태학적, 생화학적 및 배양학적 실험과 Bergey's Manual of systematic Bacteriology의 색인을 통하여 Streptomyces lydicus로 동정하였다. 효소 생산의 최적 조건을 검토하여 본 결과 배양 후 6일이 경과하였을 때, 배지의 초기 pH는 8, 배양 온도는 $30^{\circ}C$, 배지에 2의 chitin을 첨가하여 배양하였을때 효소가 가장 많이 생산되었다. 또한 soluble starch, dextrin을 탄소원으로 하여 배양하였을 때, ammonium oxalazte, tryptone, casamino acid를 질소원으로 하여 배양하였을 때 효소 생산이 높게 일어났다.

• 한국미생물·생명공학회지
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• 제29권3호
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• pp.142-148
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• 2001

경주 인근 지역의 토양으로부터 식물병원성 진균 Fusarium oxysprum과 phytophthora capsici를 동시에 길항할수있는 항진균성 항생물질 생산성 생물방제균을 분리하고, 성분을 함유한 병원진균의 세포벽을 분해하는 chitinase 생산성이 우수한 균주를 분리하고자 하였다. 분리된 생물방제균의 형태학적, 생화학적 및 배양학적으로 동정하여 잠정적으로 Bacillus amyloliquefaciens 7079로 동정하였다. 이 생물방제균이 생산하는 chitinase 생산의 최적 조건을 검토하여 본 결과 Chitin-yeast extract 배지(0.7% $K_2$$HPO_4$ 0.2% $KH_2$$PO_4$ 0.1%($NH_4$)$_2$$SO_4$ 0.05% sodium citrate 0.01% MgSO$_4$$7H_2$O 0.1% yeast extract, 0.1% coloida chitin)에서 pH는 7.0 배양온도는 3$0^{\circ}C$였고 배양한 후 3일 이 되었을 때 가장 많은 chitinase를 생산하였다. 또한 0.1% colloida chitin을 탄소원으로 하여 배양하였을 때 chitinase 생산성이 가장 좋았으며 0.15% proteose peptone NO .3 또는 0.1% tryptone 을 질소원으로 하여 배양하였을 때 효소 생산이 높게 나타났다. 선발된 생물방제균의 고추를 기주식물로한 in vivo pot 시험 결과 고추역병균 Phytophthora capsici에 좋은 길항력을 확인할 수 있었다.

• 한국식품과학회지
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• 제35권6호
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• pp.1188-1192
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• 2003

새우나 게 껍질로부터 chitin을 유일한 탄소원으로 하는 집적배양에 의하여 chitinase 활성이 우수한 균주를 2종 분리하였다. 분리균주는 형태학적, 그람염색, 16s rDNA 서열분석을 통하여 Arthrobacter nicotianae로 동정되어, 각각 Arthrobacter nicotianae CH4와 Arthrobacter nicotianae CH13으로 명명하였다. 두 종의 분리균주로부터의 chitinase는 모두 pH $3.0{\sim}9.0$에서 90% 이상의 효소활성을 유지하여 높은 pH안정성을 나타내었다. 온도의 영향은 $20{\sim}60^{\circ}C$ 구간에서 최적 효소 활성의 $70{\sim}90%$를 유지하여 열안정성도 뛰어났다. A. nicotianae CH4와 A. nicotianae CH13이 분비하는 chitinase 조효소를 0.1% colloidal chitin 기질에 반응시켜서 반응산물로 생산되는 chitin 올리고당을 HPLC를 사용하여 분석하였다. A. nicotianae CH4 조효소에 의한 효소반응산물로는 올리고머인 $(GlcNAc)_4$이, A. nicotianae CH13는 단량체인 $(GlcNAc)_1$이 전체 반응산물의 각각 98% 이상 생성되었다.

• 한국미생물·생명공학회지
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• 제47권1호
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• pp.96-104
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• 2019

Beauveria bassiana, one of the most common entomopathogenic fungi, has been isolated, pre defined and characterized in-house from soil of tea cultivation area. Experiments have been performed to verify the presence of chitinase as intracellular metabolite and its release as extracellular product rendering the spores with biopesticide activity. Although there are many responsible enzymes for the pest killer action of B. bassiana, binding property of chitinase depending on presence as well as absence of serine supplemented in the media has been studied with respect to the production and kinetics. A programmed investigation conclusively indicates that the isolated spore (hyphae) of B. bassiana has been metabolically enriched with the enzyme chitinase in presence of an externally added amino acid serine with its inhibitory kinetics.

• Journal of Microbiology and Biotechnology
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• 제20권6호
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• pp.1001-1005
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• 2010

The chitinase-producing strain TKU008 was isolated from soil in Taiwan, and it was identified as a new species of Pseudomonas. The culture condition suitable for production of chitinase was found to be shaking at $30^{\circ}C$ for 4 days in 100 ml of medium containing 1% shrimp and crab shell powder, 0.1% $K_2HPO_4$, and 0.05% $MgSO_4{\cdot}7H_2O$ (pH 7). The TKU008 chitinase was suppressed by the simultaneously existing protease, which also showed the maximum activity at the fourth day of incubation. The molecular mass of the chitinase was estimated to be 40 kDa by SDS-PAGE. The optimum pH, optimum temperature, pH stability, and thermal stability of the chitinase were pH 7, $50^{\circ}C$, pH 6-7, and <$50^{\circ}C$, respectively. The chitinase was completely inhibited by $Mn^{2+}$ and $Cu^{2+}$. The results of peptide mass mapping showed that 11 tryptic peptides of the chitinase were identical to the chitinase CW from Bacillus cereus (GenBank Accession No. gi 45827175) with a 32% sequence coverage.

• 한국미생물·생명공학회지
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• 제20권5호
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• pp.511-518
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• 1992

Serratia marcescens ATCC 27117에서 부터 클로닝한 58KD 키티나아제 유전자를 subcloning 하여 2.6Kb DNA 삽입단편을 가진 플라스미드 pCHI26을 제조하고 대장균에서 발현과 분비를 살펴보았다. 키티니아제 유전자는 대장균에서 자신의 prmoter를 이용하여 매우 낮은 수준(<5mU/m$\ell$)으로 발현되었으며 lac promoter를 이용하는 경우 키티니아제 발현이 증가되어 약 80mU/m$\ell$가 되었다. 발현된 키티니아제는 거의 전적으로 대장균의 periplasm에 위치(약 87.8)하고 있었다. 배양시간에 따라서 세포내 키티니아제 활성을 측정해본 결과 초기정지기까지는 균체 성장과 비례해서 세포내 효소활성이 증가되었으나 정지기부터 세포내 효소활성이 급격히 줄어드는 양상을 보였다. 그러나 이 기간 동안 세포의 효소활성의 변화는 거의 없었다. 이러한 결과로 보아 periplasm에 위치한 키티나아제가 대장균의 단백분해효소에 의해서 분해되는 것으로 추정되었다.

• Biotechnology and Bioprocess Engineering:BBE
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• 제4권1호
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• pp.26-31
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• 1999

The advantages of the organism Streptomycs griseus HUT 6037 is that the chitinase and chitosanase using chitinaceouse substrate are capable of hydrolyzing both amorphous and crystalline chitin and chitosan. We attempted to investigate the optimization of induction protocol for high-level production and secretion of chitosanase and the influence of chitin and partially deacetylated chitosan sources (75∼99% deacetylation). The maximum specific activity or chitinase has been found at 5 days cultivation with the 48 hours induction time using colloidal chitin as a carbon source. To investigate characteristic of chitosan activity according to substrate, we used chitosan with various degree of deacetylation as a carbon source and found that this strain accumulates chitosanase in the culture medium using chitosanaceous substrates rather than chitinaceous substrates. The highest chitosanase activity was also presented on 4 days with 99% deacetylated chitosan. The partially 53% deacetylated chitosan can secrete both chitinase and chitosanase which was defined as a soluble chitosan. The specific activities of chitinase and chitosanase were 0.89 at 3 days and 1.33 U/mg protein at 5 days, respectively. It indicate that chitosanase obtained from S. griseus HUT 6037 can hydrolyze GlcNAc-GlcN and GlcN-GlcN linkages by exo-splitting manner. This activity increased with increasing degree of deacetylation of chitosan. It is the first attempted to investigate the effects of chitosanase on various degrees of deacetylations of chitosan by S. griseus HUT 6037. The highest specific activity of chitosanase was obtained with 99% deacetylated chitosan.

• The Plant Pathology Journal
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• 제22권2호
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• pp.107-114
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• 2006

The white cottony stem rot pathogen Sclerotinia scierotiorum was subjected to 70 different isolates of actinomycetes indigenous to Jordan as biological control agents. Forty of them demonstrated chitinase activity on crab shell chitin agay (CCA) media and they were segregated into three groups: 14 highly active, 12 moderately active, and 14 with low activity, with average clearing zones of (4.7-8.3), (3.7-4.3), and (2.3-3.3) mm surrounding colonies on CCA, respectively. Further, these isolates were able to inhibit radial mycelium growth of the pathogen and were categorized into three antagonistic groups: 13 strong, 13 moderate, and 14 weak antagonists, with antibiosis inhibition Bones of (32.0-45.7), (22.7-31.3), and (3.7-22.3) mm, respectively. High levels of chitinase activity of the isolates Ma3 (8.3 mm), Jul (7.7 mm), and Sa8 (7.7 mm) with their antagonistic activity against mycelium growth of 45.7, 44.3, and 40.7 mm were observed, respectively. These isolates exhibited fungicidal activity against sclevotia of S. sclerotiorum. On the other hand, isolates Na5, Aj3, and Aj2 that produced no chitinase showed fungistatic effect only.