• 대한미생물학회지
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• 제22권2호
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• pp.109-116
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• 1987

As a process to establish an effective preventive measure of V. vulnificus septicemia, bactericidal effect of distilled water against V. vulnificus was studied. When about $2.0{\times}10^7\;CFU/ml$ of V. vulnificus was inoculated in distilled water, a dramatic decrease in the number of viable bacteria by 5 to $6LOG_{10}$ was observed in 5 minutes. Bactericidal kinetic curves could be divided into the first rapid killing phase until 1 minute and the later slow killing phase after then, showing the heterogeneity of the bacterial population inoculated. When V. vulnificus was inoculated in saline solutions having various salinities, significant decrease in the number of viable bacteria was noted only at salinities under 0.2%. The higher was the concentration of NaCl, the greater was the degree of protection against osmotic shock. When glucose, NaCl, $MgCl_2$, and $CaCl_2$ were diluted with deionized water to give same osmolarities and V. vulnificus was inoculated in each of them to compare the bactericidal curves plotted during the first 5 minutes after inoculation, the protection efficiencies were in the order of $MgCl_2>CaCl_2{\gg}NaCl{\gg}glucose$. Above results indicate that treatment(or thorough washing) of contaminated sea animals or other products with distilled water can be used as a preventive measure of V. vulnificus septicemia, and divalent cations can protect V. vulnifcus to osmotic shock with high efficiency.

• 한국미생물·생명공학회지
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• 제14권2호
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• pp.175-179
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• 1986

Cellulose분해력이 높은 Cellulomonas flavigena NCIB 12901의 원형질체 융합을 위한 원형질체의 형성조건을 조사하여 본 결과 배양기간에 따라 원형질체의 형성율이 매우 달라져 대수증식기 말기보다는 중기때의 세포를 lysozyme처리하는 것이 더 효율적이며 lysozyme 400$\mu\textrm{g}$/$m{\ell}$ 농도로 6시간처리 하여 95% 이상이 형성됨을 관찰할 수 있었다. 또한 원형질체 형성확인법으로 osmotic shock에 의한 원형질체 계수법은 spheroplast의 형성과 원형질체의 형성이 구별되어 지지 않으므로 그 결과가 전자현미경상으로 직접 관찰된 결과와 일치하지 않는 경우가 있어 전자현미경으로의 관찰이 뒤따라야 하는 것을 알 수 있었다.

• Journal of Microbiology and Biotechnology
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• 제25권7호
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• pp.1056-1069
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• 2015

Osmotic pressure is a critical factor for erythritol production with osmophilic yeast. Protein expression patterns of an erythritol-producing yeast, Yarrowia lipolytica, were analyzed to identify differentially-expressed proteins in response to osmotic pressure. In order to analyze intracellular protein levels quantitatively, two-dimensional gel electrophoresis was performed to separate and visualize the differential expression of the intracellular proteins extracted from Y. lipolytica cultured under low (3.17 osmol/kg) and high (4.21 osmol/kg) osmotic pressures. Proteomic analyses allowed identification of 54 differentially-expressed proteins among the proteins distributed in the range of pI 3-10 and 14.4-97.4 kDa molecular mass between the osmotic stress conditions. Remarkably, the main proteins were involved in the pathway of energy, metabolism, cell rescue, and stress response. The expression of such enzymes related to protein and nucleotide biosynthesis was inhibited drastically, reflecting the growth arrest of Y. lipolytica under hyperosmotic stress. The improvement of erythritol production under high osmotic stress was due to the significant induction of a range of crucial enzymes related to polyols biosynthesis, such as transketolase and triosephosphate isomerase, and the osmotic stress responsive proteins like pyridoxine-4-dehydrogenase and the AKRs family. The polyols biosynthesis was really related to an osmotic response and a protection mechanism against hyperosmotic stress in Y. lipolytica. Additionally, the high osmotic stress could also induce other cell stress responses as with heat shock and oxidation stress responses, and these responsive proteins, such as the HSPs family, catalase T, and superoxide dismutase, also had drastically increased expression levels under hyperosmotic pressure.

• Journal of Pharmaceutical Investigation
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• 제32권3호
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• pp.181-184
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• 2002

This study reports the encapsulation of plasmid DNA in erythrocyte ghosts. The plasmid DNA was encapsulated into erythrocyte ghosts using three methods; osmotic shock, electroporation in isotonic medium, and e1ectroporation in hypotonic medium. Of three methods, electroporation in hypotonic medium resulted in the highest encapsulation efficiency of plasmid DNA. The morphology of erythrocyte ghosts prepared by electroporation in hypotonic medium was similar to that by osmotic shock alone. The circulation time of plasmid DNA in mice was prolonged by administration in erythrocyte ghost-encapsulated forms. These results indicated the potential of erythrocyte ghosts for biocompative nonviral delivery system of therapeutic genes for hematological diseases.

• Journal of Microbiology and Biotechnology
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• 제12권5호
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• pp.841-843
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• 2002

Pfu DNA polymerase from Pyrococcus furiosus was expressed in the E. coli periplasm, and the fully active polymerase was partially purified by applying osmotic shock, ammonium sulfate precipitation, and heat treatment. This method represents a new way of expressing and purifying functional Pfu DNA polymerase without the use of chromatography.

• Journal of Microbiology and Biotechnology
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• 제14권2호
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• pp.231-236
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• 2004

The inexpensive large-scale production of pure PGA (Penicillin G Acylase) has been a commercial goal. PGA has been used as a model enzyme in the development of simple one-step purification methods. In this study, the purification of poly-His tagged PGA protein secreted into the periplasmic space was carried out by using immobilized metal-ion affinity chromatography (IMAC). The PGA gene was obtained from E. coli ATCC 11105. Codons encoding histidines were fused at the C-terminus of the PGA gene by PCR. E. coli JM109 harboring pPGA-HIS6 vector produced active his-tagged acylases in the presence of lac promoter during cultivation at $26^{\circ}C$. The maximum specific activity of the acylase purified by using one-step chromatography after osmotic shock was 38.5 U/mg and was recovered with the yield of 70％. Both 23 kDa ($\alpha$) and 62 kDa ($\beta$) subunits were recovered by using IMAC with just C-terminus tagging of the $\beta$ subunit. The purification of the periplasmic fraction by osmotic shock and that of purified acylase was increased by 2.6-fold and 19-fold, respectively, compared to the crude extract.

• 농업과학연구
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• 제32권1호
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• pp.71-80
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• 2005

본 연구에서는 phytic acid가 금속이온이나 양이온을 chelate시키는 작용이 탁월하며, 특히 $Fe^{2+}$나 $Ca^{2+}$에 대해서 친화성이 높다는 점을 착안하여, 사람이 평소에 쉽게 섭취하고 식용이 가능하며 인체에 무해한 장점이 있는PA가 EDTA를 대체할 수 있는지 여부를 알아보았고, 실제 동물 실험을 통해 생존율 효과에 얼마나 영향이 미치는지를 관찰한 결과이다. 1. 살균적 Osmotic Shock 및 Chelating Agent의 강화효과 1) 초기 실험에 투입된 균수를 측정한 결과 $1.7{\times}10^6cfu/ml$ 이었으며 증류수에 의한 osmotic shock을 받은 균의 생존균수는 $Mg^{2+}$ 경우 접종 후 1분에서 92.5%, 3분에서 91.8%, 5분에서 79.8%로 평균 88%로 큰 차이 없는 살균효과를 나타났다. 2) 증류수에 의한 osmotic shock을 받은 균은 접종 후 1분에서 83.5%, 3분에서 70.0%, 5분에서 60.2%로 평균 71.2%로 비교적 완만한 살균효과를 나타났다. 3) EDTA 및 PA 용액은 0.1mM 농도의 경우 보다 1mM 농도에서 osmotic shock가 극적으로 강화되는 경향을 알 수 있었다. 2. Phytic acid의 Vibrio vulnificus 패혈증 마우스 생존율에 미치는 영향 V. vulnificus 패혈증에 미치는 영향을 관찰한 결과 고농도의 경우 마우스의 평균 사망시간은 348분이였으나 저농도인 경우 마우스의 평균 사망시간이 1246분으로 고농도 보다 3.6배나 오랜시간 생존한 것으로 나타났다. CaEDTA의 처치군 고농도의 경우 평균 생존시간은 387분으로 대조군 348분으로 약간의 생존시간 증가의 경향을 관찰 할 수 있었고 저 농도의 경우 평균 생존시간은 669분으로 고농도에 비해 생존시간이 1.8배 정도 증가하는 경향을 관찰할 수 있었다. 그러나 phytic acid농도의 영향은 있지만 저농도에서는 생존시간이 훨씬 늘어났다. 따라서, 마우스의 접종실험결과 병원균인 V. vulnificus의 접종농도와 마우스의 치사속도가 정의 상관관계가 있는 것으로 나타났다. 3. phytic acid가 간이 손상된 마우스의 Vibrio vulnificus 패혈증에 미치는 영향 사염화탄소로 간을 파괴시킨 마우스를 사용하여 정상 마우스(대조군)와 비교한 결과, 간 손상이 없는 대조군에 비해 1.5배 빠른 사망 속도로 나타나, 기저질환인 간장질환 등을 앓고 있는 사람이 V. vulnificus에 감염될 경우 치사 속도가 치명적임을 간접적으로 관찰할 수 있었다. 마우스에 V. vulnificus의 접종량을 달리한 결과 $2.3{\times}10^7cfu/ml$에서는 접종 후 평균 생존시간은 226분이었고, $0.8{\times}10^6cfu/ml$에서는 접종 후 평균 생존시간은 300분으로 두 농도간 사망속도는 1.3배로 죽어가는 양상은 비슷하였으나, 기저 질환이 없을지라도 치사속도는 접종량에 크게 의존하고 있음을 알 수 있었다.

• 미생물학회지
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• 제24권2호
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• pp.154-160
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• 1986

Cellulose분해력이 높은 Cellulomonas속 세균의 원형질체 융합을 위한 원형질체 형성조건을 조사하여 본 결과 같은 Cellulomonas 속인 Cellulomonas sp. C S 1- 1과 Cellulomonas flavigena NCIB 12901 균주일지라도 그 형성조건에 현저한 차이가 있어 CS1-l균주는 lysozyme농도 $100{\mu}g/ml$ 30분정도의 처리로 99.9%의 높은 원형질체 형성을 얻을 수 있는 반면 C.flavigena 균주는 lysozyme 농도도 훨씬 높고 장시간이 소요되어 $600{\mu}g/ml$. 6시간 정도의 처리에 약80%의 원형질체 형성을 보였다. 또한 세균 배양기간이 미치는 영향도 CS1-l균주는 별 영향을 받지 않고 원형질체화 되었으나 C. flavigena균주는 매우 민감하게 영향을 받아 대수증식기 중기의 세포가 말기의 세포보다 원형 질체가 잘 이루어 졌다. 원형질체 형성확인방법에서도 CS-1 균주는 osmotic shock을 주어 원형질체를 계수하거나 SEM으로 확인하거나 같은 결과를 얻었으나 C. flavigena 균주는 osmotic shock에 의한 원형질체의 계수결과와 SEM으로의 결과가 서로 달라 확인방법으로 두 방법이 병행되어져야 함을 보여주었다.

• 한국생물공학회:학술대회논문집
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• 한국생물공학회 2000년도 춘계학술발표대회
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• pp.355-356
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• 2000

Three 'stress probe' plasmids were constructed and characterized which utilize a green fluorescent protein (CFP) as a non-invasive reporter to elucidate Escherichia coli cellular stress responses in quiescent or 'resting' cells. Facile detection of cellular stress levels was achieved by fusion of three heat shock stress protein promoter elements, those of the heat shock transcription factor ${\sigma}^{32}$, pretense subunit ClpB, and chaperone DnaK, to the reporter gene $gfp_{uv}$. When perturbed by chemical or physical stress (such as heat shock, nutrient (amino acid) limitation, addition of IPTG, acetic acid, ethanol, phenol, antifoam, and salt (osmotic shock), the E. coli cells produced GFPuv which was easily detected from within the cells as emitted green fluorescence. A temporal and amplitudinal mapping of these responses was performed, demonstrating regions where quantitative delineation of cell stress was afforded.

• Genomics & Informatics
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• 제7권1호
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• pp.26-31
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• 2009

Heat shock proteins are a class of molecular chaperones that can be found in nearly all organisms from Bacteria, Archaea and Eukarya domains. Heat shock proteins experience increased transcription during periods of heat induced osmotic stress and are involved in protein disaggregation and refolding as part of a cell's danger signaling cascade. Heat shock protein, Hsp20 is a small molecular chaperone that is approximately 20kDa in weight and is hypothesized to prevent aggregation and denaturation. Hsp20 can be found in several strains of Proteobacteria, which comprises the largest phyla of the Bacteria domain and also contains several medically significant bacterial strains. Genomic analyses were performed to determine a common evolutionary pattern among Hsp20 sequences in Proteobacteria. It was found that Hsp20 shared a common ancestor within and among the five subclasses of Proteobacteria. This is readily apparent from the amount of sequence similarities within and between Hsp20 protein sequences as well as phylogenetic analysis of sequences from proteobacterial and non-proteobacterial species.