• Journal of Microbiology and Biotechnology
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• 제16권8호
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• pp.1185-1191
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• 2006

The expression of a pullulanase gene in Pichia pastoris was investigated. The gene encoding pullulanase was cloned by PCR using the chromosomal DNA of Bacillus naganoensis as the template. The expression vector pPIC9K-Pu was constructed by inserting the pullulanase gene into plasmid pPIC9K and then transformed into Pichia pastoris SMD 1168 by electroporation. Activity determination, SDS-PAGE, and PCR amplification indicated that the gene of the pullulanase from B. naganoensis had successfully been expressed in SMD 1168 and the molecular size of the expressed recombinant product was about 119.9 kDa. This is the first report on the successful expression of the pullulanase from B. naganoensis in P. pastoris. The transformant secreted recombinant pullulanase with the activity of 350.8 IU/ml in shake-flask culture. The properties of the recombinant pullulanase were characterized.

• 한국생물공학회:학술대회논문집
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• 한국생물공학회 2000년도 추계학술발표대회 및 bio-venture fair
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• pp.669-671
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• 2000

Antimicrobial cationic peptides have attracted increasing research and clinical interest as a natural antibiotics due to their broad spectrum of antimicrobial activites and the rapid development of multidrug-resistant pathogenic microorganisms. In this study, first, we synthesized artificial fusion partner and cationic peptide genes (lactoferricin, magainin, protegrin-1, and indolicidin). Second, we constructed recombinant expression vectors and then transformed Pichia pastoris. Finally, expressed cationic peptides were purified and tested for their antimicrobial activites. Antimicrobial activity has been tested upon the appearance of clearing zone on the plate with the lawn of gram negative E.coli XL- I blue and garm positive Staphylococcus aureus. Protegrin-1 and Indolicidin have apparant activity of cationic peotides. This fusion technique may lead to a general and suitable tool for production of pure antimicrobial cationic peptides in Pichia pastoris.

• 한국식품영양과학회지
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• 제26권5호
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• pp.844-850
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• 1997

$\alpha$-Galactosidase was partially purified from the culture filtrate of pichia guilliermondii by Mannobiose-Sepharose affinity column chromatography. The galactosidase exhibited maximum activity at pH 4.5 and 4$0^{\circ}C$, and was stable in the pH and temperature ranges of 4 to 5.5 and 30 to 6$0^{\circ}C$, respectively. The enzyme was inhibited by $Hg^{2+}$ and $Ag^{2+}$. The enzyme activity was ot affected considerably by treatment with other metal compounds. The enzyme hydrolyzed melibiose to galactose and glucose, raffinose to galactose and sucrose, and $Gal^{3}Man_{3}(6^{3}-$\alpha$-galactosyl-1,4-mannotriose)$ to galactose and mannotriose. On the contrary, it could not hydrolyze $Gal^{3}Man_{4}(6^{3}-galactosyl-1,4-$\alpha$-mannotetraose)$.

• 한국생물공학회:학술대회논문집
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• 한국생물공학회 2002년도 생물공학의 동향 (X)
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• pp.157-160
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• 2002

생균제로 새로이 개발된 효모인 Pichia amomala를 대량 생산하기 위한 배양 실험으로 가격이 저렴하고 균주 생육에 적절한 질소원으로의 CSL 실험을 실시한 결과 Yeast extract 에 비해 보다 우수한 생육을 보였고 , CSL 을 이용한 3ton fermenter 에서의 batch 실험결과 최대 OD 58까지 배양되었고 , batch 배양시 harvest 시점은 가장 많은 viale cell 을 얻을 수 있는 시점인 배양 후 20 시간으로 결정되었다 . 또한 원심분리후 동결건조시 안정제에 따fms 동결건조시의 생존에 대한 실험을 실시한 결과 skim milk 2%, scrose 2% 에서 가장많은 생균수를 얻을 수 있었다.

• Journal of Microbiology and Biotechnology
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• 제14권1호
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• pp.68-73
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• 2004

Ferritin is an iron storage protein found in most living organisms. For expression and industrial use, human light chain ferritin (L-ferritin) was cloned from human liver cDNA library and expressed in Pichia pastoris strain GS115. The recombinant L-ferritin in Pichia pastoris was glycosylated. In a fed-batch culture, the cell mass reached about 57 g/l of dry cell weight, and the L-ferritin in the cell was increased to about 95 mg/l after 150 h. In an atomic absorption spectrometry analysis, the intracellular content of iron in the L-ferritin transformant was measured as $1,694{\pm}85\;\mu\textrm{g}g/g$, which is 5.4-fold more than that of the control strain. This L-ferritin transformant could serve as iron-fortified nutrients in animal feed stock.

• KSBB Journal
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• 제14권4호
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• pp.482-489
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• 1999

As host for the production of eucaryotic heterologous proteins, methylotrophic yeast Pichia pastoris and Hansenula polymorpha are the most highly developed of a small group of alternative yeast species chosen for their perceived advantages. This paper describes the method to enhance the recombinant protein productivity with P. pastoris and H. Plymorpha. In these experiments, the effects of methanol induction timing, induction method, pH, culture temperature and kinds of nitrogen sources on foreign protein production were tested with P. pastoris and compared with H. polymorpha.. In addition, optimum methanol concentration as inducer and the effects of carbon sources on AOX1 or MOX promoter repression and secretion efficiency were also studied in both cases.

• Journal of Applied Biological Chemistry
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• 제50권1호
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• pp.29-31
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• 2007

• 한국미생물·생명공학회지
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• 제44권3호
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• pp.349-354
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• 2016

누룩으로부터 44℃에서 성장할 수 있는 효모 균주를 분리하고 상대적으로 성장속도가 가장 우수한 MBY/L1569 균주를 선별하였다. MBY/L1569 균주는 Pichia farinosa로 동정되었으며, 대조구인 P. farinosa KCTC27412 균주에 비하여 내열성, 내산성 및 내알칼리성이 월등히 우수하였다. P. farinosa MBY/L1569 균주는 46℃에서 0.37 ± 0.05 h⁻¹의 비성장속도를 나타내었으며, 배양온도 37℃, 초기 pH 7.0, 포도당 농도 50 g/l 조건에서 0.50 ± 0.02 h⁻¹의 가장 빠른 비성장속도를 나타내었다. 최적 호기적 배양조건에서 P. farinosa MBY/L1569 균주는 50 g/l 포도당으로부터 19.66 ± 0.68 g/l 에탄올을 생산하여 약 40%의 수율을 나타냈다. P. farinosa MBY/L1569 균주는 한국미생물자원센터에 KCTC27753 균주로 기탁하였다.

• Journal of Applied Biological Chemistry
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• 제52권4호
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• pp.163-169
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• 2009

A total of 155 microbial strains were isolated from the Korean traditional soybean paste based on their morphological features on the growth of agar plate. Among the isolated strains, a total of 28 strains were capable of hydrolyzing isoflavone glycoside to isoflavone aglycone efficiently in the soybean paste. Finally, two strains, K123-1 and SI, were selected because of their resistance to 15% NaCl and ability to convert isoflavone glycoside to isoflavone aglycone efficiently during the fermentation of soybean paste. The isolated strains K123-1 and SI were identified to be Pichia guilliermondii and Candida fermentati, respectively, using the partial 26S rDNA sequence analysis and phylogenic analysis. Pichia guilliermondii K123-1 and Candida fermentati SI converted daidzin to daidzein up to 96% and 95%, respectively, and genistin to genistein up to 92% when soybean pastes were fermented at $30^{\circ}C$ for 20 days with a single isolated strain. Pichia guilliermondii K123-1 and Candida fermentati SI were able to grow in the presence of 15% NaCl on both liquid medium and agar plate. We think that Pichia guilliermondii K123-1 and Candida fermentati SI might be one of good candidates for making functional soybean paste because they are isolated from the Korean traditional soybean paste and have a good ability to convert isoflavone glycosides to isoflavone aglycones and a high salt tolerance.

• 한국식품영양과학회지
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• 제36권8호
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• pp.1077-1082
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• 2007

본 연구에서는 발효식품의 저장기간을 연장하거나 이상발효를 방지하기 위해 미생물 유래의 천연 항균성 물질인 killer toxin 생산 균주인 K3, K5, K11, K12, K15를 전통누룩으로부터 분리하였다. 분리된 killer toxin 생산 균주 중 식중독의 원인균인 Salmonella Typhimurium 및 장염비브리오의 원인균인 Vibrio parahaemolyticus의 생육을 저해하며, killer toxin 활성이 가장 우수한 K15를 최종 선발하고 이를 Biolog사 동정시스템과 ITS영역의 염기서열 homology를 조사하여 동정한 결과, Pichia anomala에 99% 상동성을 나타내어 Pichia anomala K15로 명명하였다. P. anomala K15가 생산하는 killer toxin은 단백질 분해효소에 의해 불활성화 되므로 인체에서 단백질 분해효소에 의해 쉽게 분해가 가능한 안전한 항균물질임을 확인할 수 있었다. 또한 p. anomala K15는 에탄올 내성은 약하나 고농도의 당에서 저항성이 크므로 주조 발효초기 환경에서의 이상발효를 방지할 수 있을 것으로 사료되어진다.