• 한국식품영양학회지
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• 제1권1호
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• pp.25-32
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• 1988

Properties of purified isocitrate lyase from Saccharomycopsis lipolytica ATCC 44601 and MX9-11RX8 temperature-sensitive mutant were investigated. Purified isocitrate lyase from temperature-sensitive mutant was indistinguishable from the wild type enzyme with respect to the isoelectric pH(5.3), the thermostability and Km value for threo-Ds-Isocitrate(about 0.2 mM). When isocitrate lyase induced by acetate minimal medium at 33$^{\circ}C$, MX9-11RX8 mutant did not express enzyme activity but did synthesize polypeptide chain whose electrophoretic mobilities were equal to those of the purified enzymes.

• 한국환경성돌연변이발암원학회지
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• 제10권2호
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• pp.93-106
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• 1990

The Drosophila temperature-sensitive mutant shibire (shi) is paralyzed at restrictive temperature by a reversible block in synaptic transmission. To explore the functional relationship among shi gene products, viability and temperature-sensitive paralytic behavior were quantitaively analyzed for four shi alleles, shi$^{ts1}$, shi$^{ts2}$, shi$^{ts4}$, and shi$^{ST139}$, and their heteroallelic combinations. The hemizygous combination of shi alleles over deficiency was not completely lethal. shi$^{ts2}$ exhibited distinctively higher viability than other alleles. A novel behavior, bang sensitivity, was also found in shi/Df(1). This bang-sensitive paralytic behavior was compared with that of the typical bang-sensitive mutant flies. Heterozygotes, shi/+, are more severe in temperature sensitivity than deficiency hemizygotes, Df(1)/+. Heteroallelic combinations of shi were less sensitive to high temperature than homozygotes. Among all allelic combinations, shi$^{ts2}$/shi$^{ts4}$ showed an unexpected extreme reduction in temperature sensitivity. The results of allelic interactions among 4 shi alleles suggest that the shi mutations examined behave as antimorphic alleles and that the gene product of shi are likely to function in multimeric forms.

• 미생물학회지
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• 제20권4호
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• pp.173-182
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• 1982

From FGSC 159 strain of Aspergillus nidulans, temperature sensitive mutants that are defective in growth and differentiation have been isolated by N-methyl-N'-nitroN-nitrosoguanidine (NTG) treatment. The optimum concentration of NTG and incubation time to get the highest mutation frequency was $100{\mu}g$ per ml and 1 hour, respectively. The survival frequency was 1%. Among the isolated mutants, five strains that were affected in early steps of differentiation were selected for further studies and named smK, smY, smB, smF, and smZ. The execution point of each mutant was determined and the growing pattern of each mutant at the restrictive temperature was observed under the microscope. Growth of mutant was arrested near at the execution point. From genetic analysis, each temperature-sensitive mutants was thought to have a single recessive gene. The genes of smK, smY, smB, smF, and smZ are linked to the chromosome VII, IV, VIII, I, and VI, respectively. It can be concluded that the genes controlling the differentiation are widely dispersed in the genome. From the results of mutant, smK, it is considered that a single gene can affect a function (functions) which act(s) at two different steps during differentiation.

• BMB Reports
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• 제30권3호
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• pp.167-172
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• 1997

The yeast Saccharomyces cerevisiae, mutant CH117, shows a drug-hypersensitivity (dhs) to cycloheximide, bleomycin, actinomycin D, 5-fluorouracil. nystatin, nigericin and several other antibiotics. CH 117 was also temperature-sensitive (ts). being unable to grow at $37^{\circ}C$ and secreted more invertase and acid phosphatase into the medium than the parent yeast. CH117 grows very slowly and the cell shape is somewhat larger and more sensitive to zymolyase than the wild type cells. Light microscopic and electron microscopic observation also revealed abnormality of the mutant cell wall. These characteristics indicate that CH117 has a defect in an essential component of the cell surface and that the cell wall which performs barrier functions has become leaky in the mutant. We screened a genomic library of wild type yeast for clones that can complement the mutation of CH117. A plasmid, pCHX1, with an insert of 3.6 kilobases (kbs) could complement the dhs and ts of CH117. Deletion and subcloning of the 3.6 kb insert showed that a gene for the complementation of mutant phenotypes was located in 1.9 kbs Puvll-Hindlll fragment.

• 한국미생물·생명공학회지
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• 제15권6호
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• pp.414-419
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• 1987

Saccharomycopsis lipolytica의 isocitrate lyase에 대한 온도감수성 변이의 특성과 이 효소의 생화학적성질을 조사하기 위한 기초연구로서 isocitrate lyase 결손변이 균주인 MX9-11(Icl-)로부터 온도감수성을 나타내는 복귀 변이균주를 3균주 분리하고 그중 활성의 회복이 가장 높은 MX9-11RX8 균주의 성질을 조사하였다. MX9-11RX8 복귀 변이균주는 탄소원으로서 초산을 사용하였을 때 온도의 증가에 따라서 isocitrate lyase의 생성량과 비생성 속도가 감소하여 33$^{\circ}C$에서는 거의 효소를 합성하지 않았으며, 온도를 23$^{\circ}C$에서 33$^{\circ}C$로 올렸을 때 효소의 합성은 바로 정지하였지만 균체증식은 계속되었다. 한편 n-hexadecane을 탄소원으로 사용하였을 때에도 온도감수성을 나타내었으며, 온도의 shift up 후에도 isocitric acid 생산은 거의 변화가 없었다.

• 생명과학회지
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• 제10권3호
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• pp.279-285
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• 2000

cAMP 수용성 단백질인 CRP는 Escherichia coli에서 대사와 관련된 유전자의 전사를 조절한다. 본 연구는 야생형과 돌연변이 CRP 단백질의 열적 안정성과 온도에 따른 단백질의 구조변화를 관찰하기 위 하여 proteolytic digestion, UV spectrophotometer, CD spectrapolarimeter 등의 방법을 사용하였다. cAMP가 없을 때에는 야생형, S83G, S128A CRP가 열적 안정성에서 큰 차이를 보이지 않았지만, cAMP가 존재할 때 야생형 CRP가 다른 돌연변이 CRP보다 열적으로 더욱 안정함을 보였다. 그리고 protease digestion 실험을 통하여 높은 온도에서 cAMP의 존재와 무관하게 돌연변이 CRP에서 단백질 의 변성으로 인한 절단된 단백질띠를 관찰할 수 있었다. 그리고 55$^{\circ}C$에서 측정한 CD 스펙트럼에서 단백 질의 2차 구조인 $\alpha$-helix 구조가 부분적으로 파괴되었음이 관찰되었다.

• 미생물학회지
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• 제54권4호
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• pp.309-319
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• 2018

이전의 연구에서 질소원이 존재하여도 페로몬을 유도하는 6개의 Schizosaccharomyces pombe 돌연변이체를 온도민감성 돌연변이체들의 저장고로부터 분리하였음이 보고된 바 있다. 본 연구에서는 이들 중 하나인 pws6 돌연변이체의 특성을 더 연구하였다. 이 돌연변이체는 영양물질에 특이적으로 페로몬 유도를 나타내었다. 즉 질소의 고갈은 없어도 M-factor 페로몬을 유도하였으나 탄소의 고갈이 없으면 유도되지 않았다. 이러한 결과는 pws6 돌연변이체가 질소 고갈을 전달하는 경로에 특이한 결함을 가지고 있음을 시사한다. 이 돌연변이체는 M-factor 페로몬뿐만 아니라 P-factor 페로몬도 온도에 민감한 양식으로 질소의 고갈 없이 유도함을 보여 주어 이 돌연변이체의 페로몬 유도는 세포 유형에 특이적이지 않음을 시사하였다. 이 돌연변이체의 온도 민감성 성장 결함의 상보적 보완에 의해 $pws6^+$ 유전자를 클로닝하여 8.1 kb, 3.3 kb, 그리고 4.8 kb 효모 DNA를 가진 3개의 플라스미드가 분리되었다. 이 플라스미드들은 pws6 돌연변이체의 성장 결함을 각각 100%, 70%, 그리고 10-20% 보완하였다. 또한 이 플라스미드들은 pws6 돌연변이체의 페로몬 유도 특성을 보완하는 능력을 가지고 있었으며 이는 돌연변이체의 성장 결함 보완 효율과 밀접한 연관성이 있음을 보여 주었다. 이들의 오픈 리딩 프레임을 성장 결함의 보완 효율과 비교하여 오픈 리딩 프레임 SPBC19C7.06이 pws6 돌연변이체의 온도 민감성 특성을 상보적으로 보완하는데 원인이 되는 리딩 프레임으로 결론 내렸다. 이 오픈 리딩 프레임은 prs1으로 명명되었으며 인트론이 없이 하나의 긴 엑손을 가지고 있는 추정된 prolyl tRNA synthetase를 암호화한다. 추정된 Prs1 단백질은 다른 종의 prolyl tRNA synthetase와 상당한 유사성을 보여 주었다.

• 한국미생물·생명공학회지
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• 제21권3호
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• pp.214-220
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• 1993

The mating type a of yeast, Saccharomyces cerevisiae mutant with hmla2-102 and sir3-8ts was changed to type alpha by changing the growth temperature from 25C to 35C. A temperature-sensitive expression vector system was constructed using mating factor alpha1 (Mfalpha1) gene encoding alpha factor which is expressed in the type alpha cells. Vectors with different copy numbers were constructed by joining the promoter and pre or prepro-secretion single sequence of Mfalpha1 to promoterless PHO5' gene as a reporter gene.

• 대한수의학회지
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• 제53권4호
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• pp.211-216
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• 2013

This study was focusing on evaluating the protection of polyphosphate kinase (ppk) deleted and/or temperature-sensitive (ts) Salmonella Enteritidis (SE) as an attenuated vaccine in chickens. We constructed SEppk, SEts and SEppk::ts mutants and screened those mutants by growth capability in vitro, protection study in mice model and antibody response in chickens. Among the mutants, SEppk::ts-3 was selected because it showed higher growth capability, good protection against highly virulent SE in mice model, and good antibody response in chickens. SEppk::ts-3 also showed good protection against highly virulent SE isolate because it decreased colonization of virulent SE challenge strain in spleen, liver and cecum compared with the non-vaccinated control. The SEppk::ts-3 mutant showed cross-protection against S. Gallinarum (SG) challenge although the its cross-protection rate was a little lower than that of SG9R, a commercial vaccine against SG infection. To use for live attenuated vaccine in chickens, it should further be characterized.

• 한국미생물생명공학회:학술대회논문집
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• 한국미생물생명공학회 2000년도 Proceedings of 2000 KSAM International Symposium and Spring Meeting
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• pp.78-89
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• 2000

Protein secretion in filamentous fungi has been shown to be restricted to actively growing hyphal tips. To determine whether an increase in the amount of growing surface area of a fungus can lead to an increase in the amount of protein secretion, we examined secretion in a temperature-sensitive Neurospora crassa mcb mutant that shows a loss of growth polarity when incubated at restrictive-temperature. Incubation of the mcb mutant at restrictive-temperature results in a three- to five-fold increase in the level of extracellular protein and a 20- fold increase in carboxymethyl cellulase activity relative to a wild-type strain. A mutation in the cr-l gene has been shown previously to suppress the apolar growth phenotype of the mcb mutant, and we find that the level of extracellular protein produced by a mcb; cr-l double mutant was reduced to that of the wild-type control. Immunolocalization of a secreted endoglucanase revealed that proteins are secreted mainly at hyphal tips in hyphae exhibiting polar growth and over the entire surface area of bulbous regions of hyphae that are produced following a shift of the mcb mutant to restrictive-temperature. These results support the hypothesis that secretion of extracellular protein by a filamentous fungus can be significantly increased by mutations that alter growth polarity.