• 한국미생물·생명공학회지
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• 제29권1호
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• pp.12-17
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• 2001

Aspergil-lus coreanus NR-15 and Aspergilus oryzae NR-2-5 from traditional Korean Nuruk were selected as parental strains producing starch hydrolysis enzyme. Xll(Arginine-) mutant from A. coreanus NR 15-1 showed high glu-doamylase activity and total acid productivity. Z6(Adenine-) mutant from A. oryzae NR2-5 showed the highest $\alpha$-amylase activity. Therefore, both XII and Z6 mutants were selected and investigated for the optimal conditions of protoplast formation for protoplast fusion. Mixture of equal amount of cellulase and driselase(10mg/ml each) was the most effective as lytic enzymes. The optimal pH and temperature for protoplast formation were 5.0 and $30^{\circ}C$, respectively. The most effective reaction for protoplast formation time was 4 hours. The maximum of protoplst for- mation of Xll mutant and Z6 mutant were $6.54$\times$10^{7}$ protoplasts/ ml and $3.04$\times$10^{ 7}$ protoplasts/ml, and the regen-eration frequencies of the protoplasts were 11.3% and 11.6%, respectively. The size of the protoplasts from X11 and Z6 mutants were 3~6 $\mu\textrm{m}$ and 4~9$\mu\textrm{m}$, respectively.

• 한국식품영양과학회지
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• 제29권6호
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• pp.987-994
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• 2000

한국 전통누룩으로부터 분리.동정된 Asp. coreanus NR 15-1, Asp. oryzae NR 15-3, 그리고 Asp. oryzae NR 2-5로부터 amylase활성이 높고 세포융합에 사용될 변이주를 분리하고 그의 특성을 검토했다. Aspergillus 속 사상균을 변이처리하기 위한 변이원으로는 NTG를 사용하였으며 Asp. coreanus NR 15-1로부터 총 15종의 변이주중 8종의 영양요구변이주, Asp. oryzae NR 15-3 으로부터 총 5종중 3종의 영양요구변이주를, Asp. oryzae NR 2-5로부터 총 6종중 2종의 영양요구변이주를 분리하였다. 이들 영양요구변이주는 최소배지 에서는 생육하지 않았으며 특정 아미노산과 핵산염기 등을 배지에 첨가했을 경우에만 생육하였다. 분리된 변이주의 당화력, 액화력 및 산 생성능 등을 검토한 결과, 산 생성능이 우수한 균주인 Asp. coreanus NR 15-1로부터 glucoamylase 활성이 높은 \ulcorner(Arg.￣) 변이주와 Asp. oryzae NR 2-5로 부터 $\alpha$-amylase 활성이 우수한 영양요구변이주인 Z6(Ade.￣) 변이주를 세포융합에 사용이 가능한 변이주라 사료되었다. 앞으로 이 들의 변이주를 대상으로 세포융합을 통해 우수한 사상균주를 획득할 수 있는 기초적 자료가 됨으로서 전통누룩의 과학화에 기여할 뿔만 아니라 외국 주류와 경쟁력이 있고 과학화된 전통주류의 개발이 가능하리 라 사료된다.

• Journal of Microbiology and Biotechnology
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• 제14권1호
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• pp.182-187
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• 2004

Strain NR $15-1^T$ isolated from traditional Korean Nuruk is described as a new species and named as Aspergillus coreanus NR $15-1^T$ sp. novo Strain NR $15-1^T$ grew rapidly to form yellow-green colonies whose surfaces were velvety on Czapek solution agar. Conidial heads were yellow to light and elliptical, whereas the conidiophore was colorless and typically long. In addition, vesicles were from flask-shaped to globose, and sterigmata are uniseriate. Conidia were spherical and deep yellow-green, and their surfaces were lightly roughened. The G＋C content of strain NR $15-1^T$ was 51 mol% and strain NR $15-1^T$contained a dihydrogenated ubiquinone with Q9 (94.9%) as a major quinone. The nucleotide sequences of strain NR $15-1^T$ in the two Internal Transcribed Spacers (ITS 1 and 2) and 5.8S rDNA showed highest similarity when compared with that of A. tubingensis and A. phoenicis NRRL $365^T$. However, based on morphological and chemotaxonomic characteristics, this strain was different from A. tubingensis and A. phoenicis NRRL $365^T$. On the basis of the data presented, it is proposed that strain NR $15-1^T$ should be placed in the genus Aspergillus as a new species, Aspergillus coreanus sp. novo Therefore, the type strain of the new species is strain NR $15-1^T$ (=KCTC 18075P^T,=KCCM 80006^T$.

• KSBB Journal
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• 제19권4호
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• pp.301-307
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• 2004

한국 전통누룩으로부터 분리한 신종 Aspergillus coreanus NR 15-1가 생산하는 a-amylase의 효소학적 특성을 조사했다. 공시균주가 생산하는 a-amylase는 황산암모늄을 이용한 분별 침전, CM-cellulose, DEAE-cellulose, Sephadex G-100, hydroxyapatite column chromatography를 통하여 8.7%의 수율을 보이며, 78배로 정제되었다. 공시균주의 a-amylase의 분자량은 Sephadex G-100 겔 여과에 의해 49 kDa으로 나타났으며, SDS-PAGE에 의하여 51 kDa으로 측정되어 본 효소는 monomer로 추정할 수 있었다. 정제효소는 pH 4.0∼11.0 사이에서 안정하였으며, 반응최적 pH는 5.0이었고, 5$0^{\circ}C$ 이하의 온도에서 비교적 안정하며, 반응최적온도는 45$^{\circ}C$로 나타났다. 정제효소는 금속이온에 의해 효소활성에 영향을 받지 않았으나, N-bromosuccinimide에 의해서는 효소활성이 완전히 저해되어 본 공시균주의 a-amylase의 활성부위에는 tryptophan 잔기가 관여한다고 추정할 수 있었다. 정제효소의 전분분해물은 maltose, maltotriose 등의 oligosaccharide를 형성하므로 a-amylase임을 확인할 수 있었다. 신종 누룩시상균인 Aspergillus coreanus NR 15-1의 a-amylase는 5$0^{\circ}C$ 이하의 온도와 pH 4.0∼11.0사이에서 안정하여 온도와 pH의 안정성이 우수하여 누룩제조용 사상균으로 사용이 가능함을 알 수 있었다.

• Journal of Microbiology and Biotechnology
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• 제15권2호
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• pp.239-246
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• 2005

Some characteristics of two forms of glucoamylase (glucan 1 A-$\alpha$-glucosidase, EC 3. 2. I. 3) purified from Aspergillus coreanus NR 15-1 were investigated. The enzymes were produced on a solid, uncooked wheat bran medium of A. coreanus NR 15-1 isolated from traditional Korean Nuruk. Two forms of glucoamylase, GA-I and GA-II, were purified to homogenity after 5.8-fold and 9.6-fold purification, respectively, judged by disc- and SDS-polyacrylamide gel electrophoresis. The molecular mass of GA-I and GA-II were estimated to be 62 kDa and 90 kDa by Sephadex G-1OO gel filtration, and 64 kDa and 91 kDa by SDS-polyacrylarnide gel electrophoresis, respectively. The optimum temperatures of GA-I and GA-II were 60$^circ$C and 65$^circ$C, respectively, and the optimum pH was 4.0. The activation energy (Ea value) of GA-I and GA-II was 11.66 kcal/mol and 12.09 kcal/mol, respectively, and the apparent Michaelis constants (K_{m}) of GA-I and GA-II for soluble starch were found to be 3.57 mg/ml and 6.25 mg/ml, respectively. Both enzymes were activated by 1 mM Mn^{2+} and Cu^{2+}, but were completely inhibited by 1 mM N­bromosuccinimide. The GA-II was weakly inhibited by 1 mM p-CMB, dithiothreitol, EDTA, and pyridoxal 5-phosphate, but GA-I was not inhibited by those compounds. Both enzymes had significant ability to digest raw wheat starch and raw rice starch, and hydrolysis rates of raw wheat starch by GA-I and GA-II were 7.8- and 7.3-fold higher than with soluble starch, respectively.