• 한국수산과학회지
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• 제30권6호
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• pp.948-955
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• 1997

Immunomagnetic separation of virus coupled with .reverse transcription-polymerase chain reaction (IMS-PCR) was performed with infectious hematopoietic necrosis virus (IHNV). A DNA fragment of expected size was synthesized in the RT-PCR with total RNA extracted from IHNV inoculated CHSE-214. In a SDS-PAGE analysis, a protein band of over 70kDa was detected from non-infected cells and cells inoculated with IHNV and infectious pancreatic necrosis virus (IPNV). This protein was detected in the Western blot analysis probably because of non-specific reaction to monoclonal antibody against IHNV nucleocapsid protein. In the immunomagnetic separation, magnetic beads coated with monoclonal antibody against the IHNV nucleocapsid protein was incubated with supernatant from IHNV inoculated CHSE-214 cells. During this process, the non-specifically reacting protein could be removed by washing the magnetic bead with PBS in the presence of an external magnetic field, and viral proteins were detected from the remaining, cleaned magnetic beads. It was necessary to extract viral RNA from the captured virus particles before RT-PCR, and no DNA product was detected when the captured virus was only heated 5 min at $95^{\circ}C$. A PCR-product of expected size was synthesized from IMS-PCR with magnetic beads double coated either by goat anti-mouse IgG antibody -monoclonal antibody or streptavidin - biotin conjugated monoclonal antibody.

• Journal of Microbiology and Biotechnology
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• 제6권4호
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• pp.225-231
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• 1996

The production of recombinant proteins in Escherichia coli often leads to the formation of an intracellular inclusion body. Key process steps that can determine the economics of large-scale protein production from inclusion bodies are fermentation, inclusion body recovery, and protein refolding. Compared with protein refolding and fermentation, inclusion body recovery has received scant research attention. Nevertheless, it can control the final product yield and hence process cost for some products. Optimal separation of inclusion bodies and cell debris can also aid subsequent operations by removing contaminant particulates that foul chromatographic resins and contain antigenic pyrogens. In this review, the properties of inclusion bodies and cellular debris are therefore examined. Attempts to optimise the centrifugal separation of inclusion bodies and debris are also discussed.

• 분석과학
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• 제19권6호
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• pp.529-534
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• 2006

SDS-PAGE를 이용한 일반적인 단백질 분리는 단백질을 분자량에 따라 분리하는 방법이며 가장 보편적이고, 간단한 방법 중의 하나이다. 본 연구는 SDS-PAGE (sodium dodecyl sulfate-polyacrylamide gel electrophoresis)를 두 번에 걸쳐 동일한 분리 원리로 이차원으로 전개하는 방법의 전기영동을 시도하여 기존의 일차원 전기영동 분석법과 비교하였으며, 그 결과 향상된 분리능이 본 연구의 이차원 전기영동법에서 확인되었다. Gel에서 분리된 단백질들은 MALDI TOF MS를 이용하여 동정하여 서로 다른 단백질임이 확인되었으며, 이러한 duplex SDS-PAGE 분리법은 상대적으로 적은 비용으로 단백질 분리능을 용이하게 향상시킬 수 있는 경제적인 분석법으로 이용될 수 있을 것이다.

• Bulletin of the Korean Chemical Society
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• 제24권9호
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• pp.1339-1344
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• 2003

Flow field-flow fractionation (FlFFF) is a technology to separate the molecules by size in an open channel. Molecules with different size have different diffusivities and are located vertically in different positions when passing through an open channel. In this study, hollow fiber membranes instead of conventional rectangular channels have been used as materials for the open channel and this change would decrease the cost of manufacturing. FlFFF is a useful technique to characterize the biopolymeric materials. Retention time, diffusion coefficients and Stokes radius of analysis can be calculated from the related simple equations. Hollow-fiber flow field-flow fractionation (HF-FlFFF) has been used for the characterization and separation of protein mixture in a phosphate buffer solution and has demonstrated the potential to be developed into a disposable FlFFF channel. The important indexes for the analytical separation are selectivity, resolution and plate height. The optimized separation condition for protein mixture of Ovalbumin, Alcohol dehydrogenase, Apoferritin and Thyroglobulin is ${\dot V}_{out}/{\dot V}_{rad}=0.65/0.85\;mL/min$.

• 한국수산과학회지
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• 제37권1호
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• pp.18-23
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• 2004

Effect of initial protein concentration on the protein removal rate was assessed for seawater aquarium using a foam separator. Protein removal rate was increased and removal efficiency was decreased with the increase of initial protein concentration. Enrichment ratio was decreased and foam generation rate was increased with the increase of initial protein concentration. Total suspended solids (TSS) removal rate was increased with the increase of initial protein concentration, and TSS removal efficiency was decreased with the increase of initial protein concentration. Turbidity removal rate and removal efficiency were increased with the increase of initial protein concentration.

• Journal of Microbiology and Biotechnology
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• 제29권1호
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• pp.141-150
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• 2019

The spatial landmark protein Bud8 plays a crucial role in bipolar budding in the budding yeast Saccharomyces cerevisiae. The unconventional yeast Yarrowia lipolytica can also bud in a bipolar pattern, but is evolutionarily distant from S. cerevisiae. It encodes the protein YALI0F12738p, which shares the highest amino acid sequence homology with S. cerevisiae Bud8, sharing a conserved transmembrane domain at the C-terminus. Therefore, we named it YlBud8. Deletion of YlBud8 in Y. lipolytica causes cellular separation defects, resulting in budded cells remaining linked with one another as cell chains or multiple buds from a single cell, which suggests that YlBud8 may play an important role in cell separation, which is distinct from the function of Bud8 in S. cerevisiae. We also show that the YlBud8-GFP fusion protein is located at the cell membrane and enriched in the bud cortex, which would be consistent with a role in the regulation of cell separation. The coiled-coil domain at the N-terminus of YlBud8 is important to the correct localization and function of YlBud8, as truncated proteins that do not contain the coiled-coil domain cannot rescue the defects observed in $Ylbud8{\Delta}$. This finding suggests that a new signaling pathway controlled by YlBud8 via regulation of cell separation may exist in Y. lipolytica.

• Bulletin of the Korean Chemical Society
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• 제32권4호
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• pp.1315-1320
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• 2011

Whey protein (WP) is a mixture of proteins, and is of high nutritional values. WP has become an important source of functional ingredients in various health-promoting foods. In this study, size-exclusion chromatography (SEC) and asymmetrical flow field-flow fractionation (AsFlFFF) were used for separation and analysis of whey proteins. It was found that a lab-prepared WP from raw milk is mostly of ${\beta}$-lactoglobulin with small amount of higher molecular weight components, while a commercial whey protein isolate (WPI) powder contains relatively larger amount of components other than ${\beta}$-lactoglobulin, including IgG and protein aggregates. Results suggest that AsFlFFF provides higher resolution for the major whey proteins than SEC in their normal operation conditions. AsFlFFF could differentiate the BSA and Albumin, despite a small difference in their molecular weights, and also was able to separate much smaller amount of aggregates from monomers. It is noted that SEC was able to show the presence of low molecular weight components other than the major whey proteins in the WP samples, which AsFlFFF could not show, probably due to the partial loss of those low molecular weight species through the membrane.

• 멤브레인
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• 제19권2호
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• pp.113-121
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• 2009

실리카 입자를 기공 형성제로 사용하여 물리적 강도와 단백질 결합용량이 높은 다공성 키토산 및 키틴 친화 막을 제조하였다. 키토산 친화 막의 BSA 단백질 결합용량은 최대 21.8mg/mL이었으며, 키틴 친화 막의 lysozyme 효소 결합용량은 최대 26.1mg/mL이었다. 제조된 다공성 키토산 및 키틴 친화 막을 사용하여 단백질 용액의 loading 유량, loading 양 및 농도 변화에 따른 BSA와 lysozyme의 친화 막 여과 크로마토그래피 분리 실험을 수행하였다. 친화 막 여과 크로마토그래피 분리 실험을 통해 얻어진 loading/washing/elution의 단계로 구성된 일련의 크로마토그램으로부터 단백질 용출량과 결합수율을 구하였다. 키토산 및 키틴 친화 막에의 BSA 및 lysozyme 단백질의 결합량과 결합수율은 loading용액의 유량이 작을수록, 주입량 및 농도가 클수록 증가하였다. 이 결과로부터 실리카 입자를 기공 형성제로 사용하여 제조된 다공성 키토산 및 키틴 막은 단백질의 대규모 여과 크로마토그래피 분리를 위한 친화 막으로서 효과적인 활용이 기대된다.

• Food Science and Biotechnology
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• 제15권2호
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• pp.189-195
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• 2006

A study was conducted to develop optimal conditions for a large-scale, sequential separation method for value-added components from egg yolk. Starting with liquid egg yolk, immunoglobulin Y (IgY), phospholipids, and neutral lipids were extracted sequentially using water, ethanol, and n-hexane. The remainder was yolk proteins. Adjusting the pH of diluted egg yolk to pH 5.0-5.2 decreased phospholipids content in the supernatant after centrifugation, which was very important for preventing clogging problem of ultrafiltration filters during the subsequent concentration step for IgY separation. Extraction of precipitants after centrifugation with four volumes of 100% ethanol once removed most of the phospholipids and the purity of phospholipids was more than 85% (wt.) after drying. The subsequent extraction of precipitant from ethanol extraction with four volumes of hexane 3 times removed neutral lipids almost completely and resulted in a high protein product with minimal lipids. The sequential separation method is considered to be advantageous for large-scale separations of many valuable components from egg yolk.

• 한국축산식품학회지
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• 제35권4호
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• pp.431-440
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• 2015

Chicken bone is not adequately utilized despite its high nutritional value and protein content. Although not a common raw material, chicken bone can be used in many different ways besides manufacturing of collagen products. In this study, a multi-step procedure was optimized to isolate chicken bone collagen for higher yield and quality for manufacture of collagen products. The chemical composition of chicken bone was 2.9% nitrogen corresponding to about 15.6% protein, 9.5% fat, 14.7% mineral and 57.5% moisture. The lowest amount of protein loss was aimed along with the separation of the highest amount of visible impurities, non-collagen proteins, minerals and fats. Treatments under optimum conditions removed 57.1% of fats and 87.5% of minerals with respect to their initial concentrations. Meanwhile, 18.6% of protein and 14.9% of hydroxyproline were lost, suggesting that a selective separation of non-collagen components and isolation of collagen were achieved. A significant part of impurities were selectively removed and over 80% of the original collagen was preserved during the treatments.