• 미생물학회지
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• 제40권4호
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• pp.348-354
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• 2004

사람의 모유에 많이 함유된 human lactoferrin(hLF)은 항균 및 항 바이러스 작용이 있는 것으로 보고되고 있다. 본 연구에서는 hLf를 메탄올자화 효모인 Pichia pastoris에 cloning하고 그 발현을 RT-PCR, Northern blotting, SDS-PAGE및 Western blotting으로 확인하였다. 그 결과 2.1 kb의 hLf 유전자가 P.pastoris의 염색체 DNA로 끼어들어가 안정적으로 hLf를 발현하였다. 이 재조합 P.pastotis로부터 hLf를 포함하는 세포 추출액을 얻어 항균 작용을 연구하였다. 발현된 재조합 hLf는 Staphylococcus aureus, Micrococcus flavus 등의 그람 양성균에 대해 강력한 항균작용을 보일 뿐만 아니라 그람 음성 동물성 병원균인 Pseudomonas fluorescens ID 9631, E. coli ATCC8739, 25922,35 등과 Salmonella typhimurium 114,115 등 다양한 균에 대해서도 강력한 항균작용을 보였다. 이는 재조합 hLf가 생물학적 활성이 있다는 것을 보여준다.

• Asian-Australasian Journal of Animal Sciences
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• 제27권4호
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• pp.574-579
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• 2014

A lysozyme gene from breast of Tibetan sheep was successfully expressed by secretion using a-factor signal sequence in the methylotrophic yeast, Pichia pastoris GS115. An expression yield and specific activity greater than 500 mg/L and 4,000 U/mg was obtained. Results at optimal pH and temperature showed recombinant lysozyme has higher lytic activity at pH 6.5 and $45^{\circ}C$. This study demonstrates the successful expression of recombinant lysozyme using the eukaryotic host organism P. pastoris paving the way for protein engineering. Additionally, this study shows the feasibility of subsequent industrial manufacture of the enzyme with this expression system together with a high purity scheme for easy high-yield purification.

• 한국생물공학회:학술대회논문집
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• 한국생물공학회 2001년도 추계학술발표대회
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• pp.333-335
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• 2001

A simple control strategy of DO-stat was introduced to the recombinant rGuamerin production process in Pichia pastoris. This induction strategy consisted of two interrelated control loops ‘by which oxygen ratio of inlet gas and methanol feeding rate was controlled. Using this control strategy, over-feeding or under-feeding of methanol could be avoided in concomitance with the efficient control of dissolved oxygen level. As a result, the cell concentration reached 130 g/L and rGuamerin expression level was 450 iu/L, which was more than 40% increased result comparing with the fed-batch process using manual control of methanol feeding rate.

• 한국생물공학회:학술대회논문집
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• 한국생물공학회 2003년도 생물공학의 동향(XIII)
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• pp.191-194
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• 2003

본 연구에서는 산업적으로 중요성이 날로 증가하고 있는 PLC의 대량 생산을 위해서 B. cereus ATCC10987에서 분리된 PLC 유전자를 pPICZa C에 삽입한 P. pastoris system을 개발하여 extracellular PLC를 생산하였다. 또한, 공업적 이용을 위해 여러 조건에서 B. cereus와 P. pastoris에서 생산된 extracellular PLC의 활성도를 측정하여 특성을 살펴보았다.

• 한국생물공학회:학술대회논문집
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• 한국생물공학회 2003년도 생물공학의 동향(XII)
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• pp.233-235
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• 2003

본 연구에서는 산업적으로 중요성이 날로 증가하고 있는 PLC의 대량 생산을 위해서 B. cereus ATCC10987에서 분리된 PLC 유전자를 $pPICZ{\alpha}$ C에 삽입한 P. pastoris system을 개발하여 extracellular PLC를 생산하였다. 또한, 여러 조건에서 B. cereus와 P. pastoris에서 생산된 extracellular PLC의 활성도를 측정하여 특성을 살펴보았다.

• 한국미생물생명공학회:학술대회논문집
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• 한국미생물생명공학회 2004년도 Annual Meeting BioExibition International Symposium
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• pp.56-59
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• 2004

• Journal of Microbiology and Biotechnology
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• 제19권9호
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• pp.966-971
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• 2009

Peroxidase from Coprinus cinereus (CiP) has attracted attention for its high specific activity and broad substrate spectrum compared with other peroxidases. In this study, the functional expression of this peroxidase was successfully achieved in the methylotrophic yeast Pichia pastoris. The expression level of CiP was increased by varying the microbial hosts and the expression promoters. Since a signal sequence, such as the alpha mating factor of Saccharomyces cerevisiae, was placed preceding the cDNA of the CiP coding gene, expressed recombinant CiP (rCiP) was secreted into the culture broth. The Mut Pichia pastoris host showed a 3-fold higher peroxidase activity, as well as 2-fold higher growth rate, compared with the $Mut^s $ Pichia pastoris host. Furthermore, the AOX1 promoter facilitated a 5-fold higher expression of rCiP than did the GAP promoter.

• 한국생물공학회:학술대회논문집
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• 한국생물공학회 2000년도 추계학술발표대회 및 bio-venture fair
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• pp.376-379
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• 2000

메탄올 자화효모인 Pichis pastoris는 메탄올에 의해 유도되는 강력한 AOX1 프로모터의 존재로 인하여 외래 단백질의 생산을 위한 가장 좋은 숙주중의 하나이다. methanol fed batch phase(MFP)동안에 메탄올의 공급은 그 메탄올이 단백질의 발현을 유도하며 또한 숙주에게 에너지원으로 쓰이기 때문에 매우 중요하다. 과량의 메탄올은 세포의 성장을 저해하며, 반면에 불충분한 메탄올의 공급은 세포를 느리게 자라게 하고 생산성도 떨어뜨린다. 본 연구는 새로운 혈소판 응집 억제제인 saxatilin의 생산성, 혹은 수율을 최대화 하기 위해서 메탄올의 공급속도와 세포의 비성장속도를 조절하였다.

• 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.

• Journal of Microbiology and Biotechnology
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• 제15권1호
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• pp.202-205
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• 2005

Our previous study showed that the overexpression of carboxypeptidase Y (CPY) of Saccharomyces cerevisiae in Escherichia coli resulted in the formation of insoluble inclusion bodies. To produce soluble CPY, we designed a novel Pichia pastoris expression system, in which the following were inserted into expression vectors: three different signal sequences derived from the mating factor a1 of S. cerevisiae, an inulinase of Kluyveromyces marxianus, and the endogenous signal sequence of CPY. The expression vector pHIL-D2-SSinul-proCPY was the most effective in the production of proCPY among the vectors examined. The purified active CPY was obtained from proCPY by treating with proteinase K, followed by QExcellose ion-exchange column chromatography.