• International Journal of Industrial Entomology and Biomaterials
• /
• 제6권1호
• /
• pp.33-44
• /
• 2003

The arylphorin was purified from the pupal haemolymph of the Chinese oak silkmoth, Antheraea pernyi, and characterized physiologically and biochemically, The protein was purified by a simple preparative polyacrylamide gel electrophoresis (PAGE) and subsequent diffusive elution. The preparation was shown to be homogeneous by 7.5% native-PAGE. The native molecular weight of arylphorin was 450 kDa with a 80 kDa single subunit, suggesting hexamer, The protein contained high amounts (18.3%) of aromatic amino acids, phenylalanine (9.7%) and tyrosine (8.6%). Therefore, the protein was identified as a kind of a storage protein referred to as an arylphorin. The protein was stained by Schiff's reagent, suggesting a glycoprotein. The protein contained 4.9% (w/w) sugar and mannose and N-acetylglucosamine were major components. Also, degradation of the protein was begun by heat treatment at 90 for 20 minutes. These results showed that the A. pernyi arylphorin in the study is hexamer associated with the six subunits consisting of a 80kDa single subunit, and is different from that of Kajiura et al. (1998) in the subunit composition.

• Journal of Microbiology
• /
• 제43권4호
• /
• pp.354-360
• /
• 2005

Heat-labile enterotoxin B subunit (LTB) of enterotoxigenic Escherichia coli (ETEC) is both a strong mucosal adjuvant and immunogen. It is a subunit vaccine candidate to be used against ETEC-induced diarrhea. It has already been expressed in several bacterial and plant systems. In order to construct yeast expressing vector for the LTB protein, the eltB gene encoding LTB was amplified from a human origin enterotoxigenic E. coli DNA by PCR. The expression plasmid pLTB83 was constructed by inserting the eltB gene into the pYES2 shuttle vector immediately downstream of the GAL1 promoter. The recombinant vector was transformed into S. cerevisiae and was then induced by galactose. The LTB protein was detected in the total soluble protein of the yeast by SDS-PAGE analysis. Quantitative ELISA showed that the maximum amount of LTB protein expressed in the yeast was approximately $1.9\%$ of the total soluble protein. Immunoblotting analysis showed the yeast-derived LTB protein was antigenically indistinguishable from bacterial LTB protein. Since the whole-recombinant yeast has been introduced as a new vaccine formulation the expression of LTB in S. cerevisiae can offer an inexpensive yet effective strategy to protect against ETEC, especially in developing countries where it is needed most.

• BMB Reports
• /
• 제43권5호
• /
• pp.325-329
• /
• 2010

Protein kinase CKII (CKII), a heterotetramer composed of two catalytic ($\alpha$ or $\alpha$') subunits and two regulatory ($\beta$) subunits, plays a critical role in cell proliferation and anti-apoptosis. Recently, capsaicin was shown to trigger apoptosis. Therefore, we examined the effect of capsaicin on CKII activity. Although capsaicin induced apoptotic death in HeLa cells, CKII activity was increased in the cytosolic fraction of HeLa cells after treatment. Capsaicin did not change the expression of the $CKII{\alpha}$ and $CKII{\beta}$ proteins. Capsaicin stimulated the catalytic activity of recombinant CKII tetramer, but not the $CKII{\alpha}$ subunit. Moreover, capsaicin enhanced the autophosphorylation of $CKII{\alpha}$ and $CKII{\beta}$. Taken together, our data suggest that capsaicin stimulates the phosphotransferase activity of CKII holoenzyme by interacting with the $CKII{\beta}$ subunit.

• 한국생물공학회:학술대회논문집
• /
• 한국생물공학회 2000년도 춘계학술발표대회
• /
• pp.219-222
• /
• 2000

Physiological changes of Escherichia coli during the fed-batch fermentation process were characterized in this study. Overall cellular protein samples prepared at the different stage of fermentation were separated by 2-dimensional gel electrophoresis (2-DE), and differently expressed 15 proteins, Phosphotransferase enzyme I, GroEL, Trigger factor, ${\beta}$ subunit of ATP synthase, Transcriptional regulator KDGR, Phosphoglycerate mutase 1, Inorganic pyrophosphatase, Serine Hydroxymethyl-transferase, ${\alpha}$ subunit of RNA polymerase, Elongation factor Tu, Elongation factor Ts, Tyrosine-tRNA ligase, DnaK suppressor protein, Transcriptional elongation factor, 30S ribosomal protein S6 were identified using matrix-assisted laser desorption / ionization time-of-flight mass spectrometry (MALDI-TOF MS). When bacterial cells grow to high cell density, and IPTG-inducible heterologous protein is produced, expression level of overall cellular proteins was decreased. According to their functions in the cell, identified proteins were classified into three groups, proteins involved in transport process, small-molecule metabolism, and synthesis and modification of macromolecules.

• 대한수의학회지
• /
• 제37권4호
• /
• pp.875-882
• /
• 1997

To produce the recombinant fibronectin-binding protein(FnBP) for development of subunit vaccine against Staphylococcus aureus. The fnbp gene was amplified from the chromosomal DNA of S aureus KNU 196 strain using the polymerase chain reaction, and cloned into pGEX-4T-2. Then, the recombinant FnBP fused with glutathione-S-transferase was produced in E coli, purified by affinity chromatography, and identified its antigenicity and immunogenicity by Western blot. The recombinant FnBP produced in this study is considered to have the same property of native FnBP purified from S aureus, and is expected to be useful as a candidate for S aureus subunit vaccine.

• 생명과학회지
• /
• 제30권7호
• /
• pp.592-597
• /
• 2020

성숙 콩[Glycine max (L.) Merr.] 종자는 약 40%의 단백질을 함유하고 있으며 아이소플라본, 사포닌, 루테인, 비타민등 다양한 기능성 성분을 함유하고 있다. 그러나, Kunitz Trypsin Inhibitor (KTI) 단백질, 렉틴 단백질, 7S α' 서버유닛 단백질이 함유되어져 있어 콩의 품질과 기능성을 저하시키고 있다. 본 연구는 콩 및 콩 제품의 품질과 기능성을 저하시키는 KTI, 렉틴 및 7S α' 서버유닛 3가지 단백질이 모두 유전적으로 결핍된 콩 계통(titilelecgy1cgy1 유전자형)을 선발하기 위하여 진행되었다. 3개의 모본(개척2호, PI506876, Le-16)을 이용한 유전집단으로부터 성숙 종자에서 KTI, 렉틴 및 7S α' 서버유닛 3가지 단백질이 모두 없는 titilelecgy1cgy1 유전자형을 가진 1개의 계통을 개발하였다. 개발된 계통은 자주색 꽃, 유한신육형, 노란종피을 가지고 있으며 초장은 58 cm로 대원콩(46 cm)보다 길었다. 백립중은 27.1 g으로 대원콩(29.0 g)보다 작았다. 본 연구를 통하여 선발된 계통은 KTI, 렉틴 및 7S α' 서버유닛 3가지 단백질이 모두 없는 고품질 기능성 콩 품종 육성을 위한 중간모본으로 이용될 수 있을 것으로 사료되었다.

• 생명과학회지
• /
• 제9권2호
• /
• pp.146-152
• /
• 1999

본 연구에서는 대장균 트립토판 중합효소(tryptophan synthase) $\alpha$ 소단위체의 폴딩에 있어 24번 잔기의 역할을 보고자 하였다. 24번 잔기인 트레오닌이 메티오닌, 알라닌, 세린, 류신 또는 리신으로 치환된 단백질를 대장균내에서 과량 발현시켜 수용성 구조의 양과 비수용성인 inclusion body양을 조사하였다. 그 결과 메티오닌이나 류신으로 치환된 $\alpha$ 소단위체는 트레오닌이 있는 소단위체와 마찬가지로 수용성 구조가 대부분을 차지하였다. 반면, 세린이나 알라닌 또는 리신으로 치환된 단백질들은 많은 양이 inclusion body로 발현되었다. 이 결과는 트립토판 중합효소 $\alpha$ 소단위체의 폴딩에 있어서 24번 잔기가 관여하고 있음을 제시하고 있다.

• Journal of Microbiology and Biotechnology
• /
• 제8권4호
• /
• pp.361-368
• /
• 1998

The E2 protein of HCV (hepatitis C virus) is thought to have a potential role in the development of subunit vaccines and diagnostics. To express it by the insect cell/baculovirus expression (Bacu) system, we constructed a recombinant Autographa californica nuclear polyhedrosis virus (AcIL3E2), determined the most appropriate expression conditions in terms of host cell line and culture medium, and characterized the expressed HCV E2 protein. A culture system using Trichoplusia ni BTI-TN5Bl-4 cells and SF 900IISFM medium expressed a relatively high level of HCV E2 protein. It was revealed that its glycosylation properties and subcellular localization were almost the same as the ones in the mammalian cell expression system previously reported, suggesting the recombinant HCV E2 protein derived from our Bacu system can be utilized for development of a subunit vaccine and diagnostics. Interestingly, HCV E2 protein was not degraded at all even at 43 h post-heat shock in the heat shock-induced necrotic cells, probably due to its integration into the microsomal membrane, indicating that heat shock can be employed to purify HCV E2 protein.

• 한국연초학회지
• /
• 제17권1호
• /
• pp.20-26
• /
• 1995

This study was carried out to obtain the basic data which include the change of the photosynthetic rate and protein content according to growth stage in the process of senescence of tobacco plant The photosynthetic rate was the maximum with 26.31$\mu$mol.CO2/m2.sec and stomatal resistance was the minimum with 0.2552cm/sec at 15th days after leaf emergence. However, after 50 days the photosynthesis was very little occurred. During leaf developments the number of chloroplast was increased and reached at the maximum at 25th days after emergence of leaf, thereafter, it was decreased gradually. The content of protein increased continuously and showed the highest value at 15th days after leaf emergence. The degradation rate of soluble protein was more rapid than that of insoluble protein at early stage of senescence. The range of decrement in the insoluble protein was low at late stage of senescence. The content of Rubisco, the key enzyme of photoamthesis, corresponded to about 50% of soluble protein and reached to the maximum at 150 days after leaf emergence. As the senescence progressed, the content of large subunit(UV) of Rubisco showed a tendency to decrease more rapidly than that of small subunit(SSU). The total amount of amino acids was the highest at 15th days after leaf emergence.

• Applied Biological Chemistry
• /
• 제20권1호
• /
• pp.26-32
• /
• 1977

대두(大豆)(Glycine max cultivar Gwang-gyo)의 각 성숙시기(成熟時期)에 나타나는 7S globulin을 분리하여 Davis 방법(方法)에 의한 전기영동과 PAWU용매에 의해서 유리(遊離)되는 그들의 subunit를 전기영동한 결과 7S globulin중의 복합단백질간(複合蛋白質間)에는 그 구성(構成) subunit에 유사성(類似性)이 있음을 시사하였다. 7S globulin의 복합단백질(複合蛋白質)을 DEAE-Sephadex A-50으로 크로마토그라피하여 분리하였다. 이때 pH 7.6의 인산완충액(燐酸緩衝液)에서 NaCl의 농도구배(濃度句配)가 0.28M부터 0.40M 사이에서 두 개의 분획(分劃)으로 분리되었다. 이들 명(名) 단백질(蛋白質)의 subunit를 5M urea와 1% SDS로 유리(遊離)시켜 7.5% acrylamide-PAWU gel과 5.6% acrylamide-SDS gel에서 전기영동하였다. 그 결과 subunit의 하전량(荷電量)에 의해서 분리되는 PAWU gel전기영동에서 7S globulin이 5개의 주 분리대로 분리되고 그중 2개의 분리대가 7S-A globulin과 7S-B globulin에 공유(共有)되어 있었다. 또 subunit의 분자량(分子量)에 따라서 분리되는 SDS gel 전기영동에서는 7S globulin이 7개의 주 분리대를 나타내는데 그 중에서 3개의 분리대가 7S-A와 7S-B 분획에 공유(共有)되어 있었다. 따라서 7S globulin의 복합단백질간(複合蛋白質間)에는 구성(構成) subunit간(間)에 유사성(類似性)이 있는 것으로 나타났다.