• Mass Spectrometry Letters
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• 제10권3호
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• pp.84-87
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• 2019

Liquid chromatography mass spectrometry is widely employed in proteomics studies. One of such instruments is the Liquid Chromatography (LC)-Matrix-assisted laser desorption ionisation (MALDI)-Time of flight (TOF) or LC-MALDI-TOF/TOF. In this study, this instrument was used to identify the membrane proteins of a protozoan parasite namely Entamoeba histolytica. It causes amoebiasis in human. The E. histolytica trophozoites were cultured prior to the membrane protein extraction using the conventional method, $ProteoPrep^{(R)}$ and $ProteoExtract^{(R)}$ kits. Then, the membrane protein extracts were trypticdigested and analysed by LC-MALDI-TOF/TOF. Approximately, 194 proteins were identified and 27.8% (54) were predicted as membrane proteins having 1 to 15 transmembrane regions and signal peptides by combining all three extraction methods. Also, this study has discovered 3 unique proteins as compared to our previous study which merit further investigation.

• 대한임상검사과학회지
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• 제51권1호
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• pp.78-85
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• 2019

막 단백질은 심장질환, 암과 같은 우리 주변에서 흔히 발생하는 질병에 관련되어 있다. 이러한 암과 같은 특정한 질환 상태에서, 막 단백질과 관련된 신호 전달의 비정상은 세포분열을 통제하지 못하고 증가시킬 수 있으며 막 단백질의 발현에 변화가 생긴다. 막 단백질은 지질 이중층으로 이루어진 소수성 환경을 가지고 있어 불안정하기 때문에 막 단백질을 추출해서 연구를 수행하는데 어려움이 있다. 이번 연구에서는 최적화된 막 단백질 추출법을 확인하고자 서로 다른 두 가지 추출법의 효율성을 평가하였다. 두 가지 방법으로, high-speed centrifuge법과 reagent법이 비교되었다. 비교 분석결과, 미토콘드리아 내막 단백질 분석에는 high-speed centrifuge법이 효율적이고, 소포체 막 단백질 분석에는 reagent법이 유용함을 확인하였다. 게다가 유전자 온톨로지 소프트웨어를 이용해서 추출된 막 단백질의 기능분석을 진행하였을 때, 유전자 온톨로지는 reagent법에서 소포체 막 단백질에 연관된 반응이 활성화 되는 것을 확인할 수 있었다. 프로세스 네트워크 분석에서, high-speed centrifuge법에서는 하나의 클러스터를 형성화는 반면, reagent법에서는 네 개의 클러스터를 형성하는 것을 시각화하여 확인하였다. 결론적으로, 두 가지 분석법은 서로 다른 하위 막 단백질의 분석에 유용함을 확인할 수 있었다. 이러한 결과를 토대로, 막 단백질을 분석할 때, 표적의 세부 막 단백질을 고려하여 방법론을 선택하는데 도움을 줄 것으로 기대된다.

• Asian-Australasian Journal of Animal Sciences
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• 제19권4호
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• pp.601-604
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• 2006

The objective of this study was to investigate technical methods for extraction of mucopolysachharide-protein containing chondroitin sulfate from keel cartilage of chickens. The chemical composition of chicken keel cartilage was determined. For the preparation of mucopolysaccharide-protein from lyophilized chicken keel cartilage, hot water extraction and alcalase hydrolysis methods were examined. Results showed that the optimum condition of hot water extraction was incubation for 120 min with a yield of 40.09% and chondroitin sulfate content of 28.46%. For alcalase hydrolysis, the most effective condition was 2% alcalase in 10 volumes of distilled water for 120 min. The yield of hydrolysate was 75.87%, and chondroitin sulfate content was 26.61%. For further separation of chondroitin sulfate from the alcalase hydrolysate, which has a higher yield than that of hot water, 60% ethanol precipitation was performed. The yield of the ethanol precipitate was 21.41% and its chondroitin sulfate content was 46.31%. The hot water extract, alcalase hydrolysate and ethanol precipitate showed similar electrophoretic migration with standard chondroitin sulfate (chondroitin sulfate A), using cellulose acetate membrane electrophoresis. These results indicated that a significant amount of mucopolysaccharide-protein containing chondroitin sulfate could be acquired form chicken keel cartilage. Therefore, keel cartilage in chicken may provide an inexpensive source of chondroitin sulfate for commercial purposes.

• KSBB Journal
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• 제9권3호
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• pp.332-338
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• 1994

역미생용액으로 단백칠을 추출하는 공정에셔 수용 액의 이온세기와 pH가 중요한 공정변수가 된다. 본 실험에셔 사용한 역미생용액은 AOT를 Isooctane에 5 50mM의 농도로 용해시켜 사용하였고 단백칠로서 alkaline protease를 model compound로 사용하여 추출설험을 수행한 결과 본 설험의 범위에서 pH가 낮을수록, 이온세기가 낮을수록 효율적인 추출이 이루어졌으며, pH가 3.0 그리고 이온세기가 O.lM KCI 일때 분배계수가 4.0으로 가장 크게 나타났다. 단백질의 용매추출 장치로서 설관막 모률을 제작 하여 alkaline protease를 역미생을 이용하여 용매 추출을 수행하였는 바, 단백질 분리공정에 효과적인 장치로 사용될 수 었음을 보여주였다. 본 실험의 조 건에서 총괄물질전달계수 Ko가 $6.7{\times}10^{-5}cm/s$이었으며 유기용매상의 개별물칠전달저항이 수용액상의 물질전달저항보다 중요하게 나타났다. 실관막 추출 장치의 조업시 설관막 양쪽 상의 압력차를 $0.1kg/cm^2$. 이하로 조절하여 emulsion의 형성을 방지할 수 있었다.

• 한국막학회:학술대회논문집
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• 한국막학회 2004년도 Proceedings of the second conference of aseanian membrane society
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• pp.179-182
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• 2004

This study reports the extraction behavior of $\alpha$-lactalbumin using bis(2-ethylhexyl) sulfosuccinate sodium (AOT) reverse micelles. Forward extraction of $\alpha$-lactalbumin in the reverse micellar organic phase from aqueous feed solutions was strongly dependent on the AOT concentration and the complete forward extraction of 0.03 mM $\alpha$-lactalbumin was successfully achieved at an AOT concentration of ca. 100 mM. A similar dependency of the forward extraction on the AOT concentration was obtained in isooctane, n-hexane, and n-octane systems. In the backward extraction from the micellar organic phase, the recovery of the protein as high as ca. 90% was obtained with pH control and/or salt addition.

• KSBB Journal
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• 제27권3호
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• pp.177-185
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• 2012

Inonotus obliquus mushroom, which is a fungus belonging to Hymenochaetaceae family, is known to grow on birth trees in colder northern climates and to be a fungal parasite that draws nutrients out of living trees rather than from the ground. For the separation of protein-bound polysaccharide (PBP) from the culture broth and mycelium of Inonotus obliquus, three well known extraction methods namely hot water, ultrasound and microwave were used. The best extraction conditions to separate the PBP (64.94 mg/g) from mycelium by microwave were found to be for 1 hour and $150^{\circ}C$. The possibility for concentration of extracted PBP solution by using membrane was also studied. The extracted PBP solution was concentrated effectively by using an ultrafiltration membrane and the molecular weight cut off (MWCO) is 30 KDa. It was observed that a concentration by the ultrafiltration membrane is essential not only for the development of clean separation technology but also for enhanced production of PBP. As a result, we have shown that PBP in the final concentrated solution showed approximately 10 times higher than that in the crude solution by application of the developed separation systems. The separation yield of PBP was about 89.79% by gel filtration of purification steps and the purified product was confirmed to be PBP by using FT-IR.

• Journal of the Korean Association of Oral and Maxillofacial Surgeons
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• 제30권3호
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• pp.186-192
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• 2004

Purpose: Human tooth proteins are highly heterogeneous, comprising diverse proteins derived from a number of genes. The attempts to identify protein for activity of tooth matrix proteins have been defied by several factors. First, the amount of proteins within teeth is very small relative to many extracellular matrix proteins of other tissues. Second, the bioassay system is tedious and needed for long time. Therefore we tried to find easy techniques, which increase the product rate, and an assay of small proteins, with which amino acid sequence is possible without additional procedures. Materials and Methods: Total protein were extracted from 300 g enamel removed teeth and 600 g teeth with 4 mol/L guanidine HCl and purified by gel chromatography. Aliquot of proteins was implanted into muscle pouches in Sprague-Dawley rats for bioassay. By SDS-PAGE and membrane blotting, molecular weight of each protein was estimated and a partial amino acid sequence was obtained. Each fraction blotted on the membrane was cut out and inserted in rat ectopic model. Results: In dissociative method, total tooth proteins were obtained 1mg/ml from enamel removed teeth and 3.5 mg/ml from teeth. In SDS-PAGE, four clear bands at the sites corresponding to 66, 40, 20 and 18 kD. Especially The 66 kD band was clearly exhibited. Amino acid sequencing from tooth could be possible using PVDF membrane blotting technique. In amino acid sequencing, 66 kD protein was identified as albumin. Conclusion: Compared with conventional method for extraction of teeth protein and bioassay of proteins, the methods in this study were easy, time-saving and more productive technique. The matured tooth proteins omitting additional procedure of mechanical removal of enamel were simply analyzed using blotted PVDF membrane. This method seems to make a contribution as a technique for bioassay and amino acid sequencing of protein.

• 생명과학회지
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• 제10권2호
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• pp.131-139
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• 2000

Lecithin-free egg yolk protein (EYP), the by-product of lecithin extraction from egg yolk, which is denatured with an organic solvent, would normally be discarded. In this study, the denatured protein was renatured with alkali, and hydrolyzed with Alcalase in order to utilize by-product. The hydrolysate was separated through a series of ultrafiltration membranes with molecular weight cut-off (MWOO) of 10, 5 and 1 kDa, and the antioxidative activities of the hydrolysates was investigated. The 5K hydrolysate, permeate from 5 kDa membrane, showed stronger antioxidative activity than 10 K and 1 K hydrolysate which were permeated from 10 kDa and 1 kDa membrane, in a linoleic acid autoxidation system. In addition, the optimum concentration of antioxidative activity for 5 K hydrolysate was 1%, and the activity was about 37% higher as compared with α-tocopherol. The synergistic effect was also increased by using the hydrolysates with α-tocopherol.

• 동아시아식생활학회지
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• 제5권2호
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• pp.63-71
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• 1995

Defatted sesame flour is the by-products obtained after oil extracting process. Although this flour has high quality and quantity of protein its use is limited only for animal feed and fertilization. Sesame seeds contain antinutrients such as oxalate, phytate and phenol compounds and these compounds lower their nutritive value. recently, ultrafiltration(UF) has been used to concentrate protein from various food sources. This study was carried out to examine the effects of UF with different membrane pore size on the components of sesame protein concentrates including antinutrients and to compare with that of conventional acid-precipitated sesame protein isolate. The protein contents of sesame protein concentrates prepared by JF using 10K, 30K, 100K were 84.2%, 82.7%, 76.4% and the protein yields were 36.44%, 34.69, 31.43% and the protein contents was 88.7% Alkali extraction process at pH 9.0 followed by UF technique reduced oxalate and phytate content. There were 85% and 94% reduction of oxalate and phytate content by UF with membrane pore size of 100K daltons, respectively. However, the content of total phenol compounds was not reduced by this method. About 99% of calcium and 50% of zinc were removed by UF with membrane of 100K daltons. total essential amino acid contents of sesame protein concentrates prepared by UF were decreased slightly when compared with acid-precipitated sesame protein concentrate.

• 치과방사선
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• 제27권2호
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• pp.63-81
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• 1997

This study was performed in order to identify changes of the plasma membrane proteins in rat submandibular gland tumors induced by 7,12-dimethylbenz[a]anthracene [DMBA] and X-irradiation. Two kinds of tumor associated membrane proteins (protein A and B) were isolated with 3 M KCl extraction from rat submandibular gland tumors induced by DMBA and X-irradiation. To identify their antigenicities, immunoelectrophoresis and double immunodiffusion was carried out with various proteins extracted from liver, heart, skin and pancreas of adult rats and from embryonic liver, heart and skin. The rabbit antisera against the protein A did not cross-react with any of the proteins extracted from the above mentioned tissues, suggesting that protein A might be tumor specific antigen. However, the rabbit antisera against protein B was precipitated with proteins extracted from the liver of adult and embryonic rats. Polyacrylamide gel electrophoresis of these two proteins (A and B) showed that protein A was a dimer with molecular weights of 69,000 and 35,000 dalton, whereas protein B was a monomer with molecular weight of 50,000 dalton.