• 한국미생물·생명공학회지
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• 제24권6호
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• pp.726-732
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• 1996

To improve the fermentation characteristics(such as starch-degradability, ethanol tolerance, sugar and high-temperature tolerance) of recombinant haploid yeast Saccharomyces diastaticus K114, hybridization technique was used. The hybridization partner was S. diastaticus 1177 which had good glucoamylase activity and fermentabi- lity. The best hybrid HH64 showed improved ethanol tolerance, sugar and high-temperature tolerance. Especia- lly, the starch-fermentability was significantly improved, since the hybrid produced 1.60% (w/v) ethanol from 4% (w/v) starch, while the recombinant haploid K114 produced 1.30% (w/v) ethanol. The optimum temperature and pH for the starch-fermentation by the hybrid HH64 was 30$\circ$C and 5, respectively. The hybrid yeast HH64 produced 7.5% (w/v) ethanol directly from 20% (w/v) starch.

• 한국미생물·생명공학회지
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• 제22권2호
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• pp.152-157
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• 1994

Inulinase of Kluyveromyces marxianus origin was produced by recombinant yeast Saccharomyces cerevisiae under the control of GAL1 promoter, to examine the expression and localization of inulinase in a repressed(galactose-free) or derepressed(galactose-containinga) medium. The inulinase gene(INU1A) was constitutively expressed at 6.7 units/ml in a repressed medium. When the cell started to utilize galactose in a derepressed medium, the INU1A gene began to be expressed, and the final expression level reached about 45 units/ml. According to be the nondenaturingPAGE analysis, inulinase produced by S. cerevisiae was found to be less glycosylated than the bakers yeast invertase. In addition, its glycosylation pattern was less heterogeneous than the K. marxianus inulinase. The supplementation of inulin or raffinose into the derepressed medium increased the cell growth rate, while the expression of INU1A was repressed. Regardless of the carbon sources examined, most of inulinase activity (more than 98%) was found in the extracellular medium, indicating excellent secretion efficiency.

• Journal of Microbiology and Biotechnology
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• 제1권4호
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• pp.266-273
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• 1991

Hirudin, a 65-amino acid protein isolated from the salivary gland of the bloodsucking leech, Hirudo medicinalis, is a potent thrombin-specific inhibitor and blocks the thrombin-mediated conversion of fibrinogen to fibrin in clot formation. We have studied the gene expression and secretion of hirudin in yeast. Saccharomyces cerevisiae. A gene coding for hirudin was synthesized based on the amino acid sequence and cloned into a yeast expression vector $YEG{\alpha}-1$ containing the ${\alpha}-mating$ factor pre-pro leader sequence and galactose-inducible promoter, GALl0. Recombinant S. cerevisiae was found to secrete biologically active hirudin into the extracellular medium. The secreted recombinant hirudin was recovered from the culture medium and purified with ultrafiltration and reverse phase high performance liquid chromatography. Approximately 1 mg of hirudin per liter was produced under suboptimal culture conditions and brought to about 90% purity in two steps of purification.

• 한국연초학회지
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• 제23권2호
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• pp.133-140
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• 2001

PAP-I cDNA was synthesized from total RNA of Phytolacca americana leaves by RT-PCR, and then subcloned to recombinant vector pBluescript II SK-. Using PCR with primers designed in our laboratory, we could get the 9 deletion mutant PAP-I cDNA fragments. The first of the fragments was deleted by 66bp from immature N-terminal and then the rest were deleted by 90bp sequentially. Sequentially deletion mutant PAP-I cDNAs were inserted to pAc55M, on down-stream of gall promoter. Recombinant pAc55M was transformed to yeast cells, psy1 and the cells were spreaded on SC_urn-/glucose plate media. Colonies on SC_ura-/glucose plate were streaked on the same position of SC_ura-/glucose and SC_ura-/galactose plate, and we selected colonies growing on both plates, which carry non-cytotoxic deleted mutant PAP-I cDNA. We selected 4 deletion mutant PAP-I cDNAs which have not cytotoxicity.

• 한국식품영양과학회지
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• 제28권2호
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• pp.348-352
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• 1999

We have intended to accelerate the secretion of human lysozyme(HLY) with permeabilizing agents from the cultivated cells of the recombinant Saccharomyces cerevisiae. The five agents CaCl2, Tween 80, ethanol, Triton X 100, and cetyltrimethylammonium bromide(CTAB) were used as permeabilizing agents. Treatments of the yeast cell with CaCl2, Tween 80, and ethanol were effective to increase the secretion from the yeast cells. Especially, treatment of 10% ethanol increased the extracellular HLY activity by 38.6% at 30oC for 48 h in culture broth. But Triton X 100 and CTAB unexpectedly didn't play a role in increase of HLY secretion. Recovery of a foreign protein by permeabilizing agents is easier than by osmotic shock, and is less expensive than enzymatic digestion.

• 미생물학회지
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• 제26권1호
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• pp.32-36
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• 1988

The yeast gene HOM3 encodes aspartokinase, which catalyses the first step (aspartate to and from beta-aspartyl phosphate) of common pathway to threonine and methionine. The yeast HOM3 gene expression is known to be regulated by threonine and methionine specific control, and also by general control of amino acid biosynthesis. Isolation and characterization of the HOM3 gene are essential for the molecular genetic study on its regulation of expression. A recombinant plasmid pSC3 (15.5kb, vector YCp50) has been cloned into E. coli HB101 from yeast genomic library through their complementing activity of HOM3 mutation in a yeast recipient strain M34-24B. Organization of the plasmid was characterized by delineation of restriction cleavage sites in the insert fragment.

• Journal of Microbiology and Biotechnology
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• 제32권11호
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• pp.1373-1381
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• 2022

Fructan is a polysaccharide composed of fructose and can be classified into several types, such as inulin, levan, and fructo-oligosaccharides, based on their linkage patterns and degree of polymerization. Owing to its structural and functional diversity, fructan has been used in various fields including prebiotics, foods and beverages, cosmetics, and pharmaceutical applications. With increasing interest in fructans, efficient and straightforward production methods have been explored. Since the 1990s, yeast cells have been employed as producers of recombinant enzymes for enzymatic conversion of fructans including fructosyltransferases derived from various microbes and plants. More recently, yeast cell factories are highlighted as efficient workhorses for fructan production by direct fermentation. In this review, recent advances and strategies for fructan biosynthesis by yeast cell factories are discussed.

• 생명과학회지
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• 제15권6호
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• pp.923-927
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• 2005

박테리오신의 일종인 Leucocin A를 생산하는 효모의 제작을 위하여 114 bp 길이의 종지코돈을 포함하는 Leucocin A 유전자를 합성하여 효모운반체인 pYDl에 클로닝하였다. 이렇게 제작된 재조합 DNA를 효모세포에 형질전환시켜 Leucocin A를 생산하는 형질전환 효모세포를 제작하였다. 형질전환 효모는 고초균(Bacillus subtilis)에 대해 항균활성을 나타냈다. 형질전환 효모로부터 분리한 플라스미드를 주형(template)으로 하고 Leucocin A에 특이적인 primer들을 이용하여 PCR 반응을 행한 결과, 효모에 도입된 Leucocin A 유전자를 확인할 수 있었다. 이 연구의 결과로 부패하기 쉬운 식품들의 보존성을 향상시키거나, 내성이 생긴 병원균의 생육을 저해하기 위한 항생제로 사용할 수 있는 박테리오신을 산업적으로 대량생산할 수 있는 효모세포를 제작하였다.

• 한국동물위생학회지
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• 제35권4호
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• pp.307-312
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• 2012

Iron deficiency anemia is also recognized as a serious disorder in many livestock, especially, piglets. We previously studied that the iron-fortified yeast (Saccharomyces cerevisiae) producing Sus scrofa ferritin heavy-chain (FER) was bioavailable to mice with iron deficiency. In this study, we determined whether FER could improve iron deficiency in piglets. The bioavailability of FER was examined by measuring body weight gain, hemoglobin concentration and hematocrit value in suckling and weaning piglets. We found that FER significantly increased hemoglobin value and the hematocrit ratio in suckling piglets (P<0.05). Furthermore, FER treatment significantly enhanced body weight gain in both groups of the suckling and weaning piglets (P<0.05). These results suggest that the iron-fortified recombinant yeast strain is helpful in iron absorption in piglets.