• 약학회지
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• 제25권4호
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• pp.137-143
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• 1981

In order to confirm the exact antipyretic component in the earthworm, etherial extract of American earthworm(Red Worm) was fractionated into five fractions by using silica gel column chromatography and thin layer chromatography. The fraction including free fatty acids was found to possess artipyretic response and standard arachidonic acid showed marked antipyretic response on typhoid vaccinated rabbits. Arachidonic acid was identified from the free fatty acid fraction of the earthworm by using gas liquid chromatography. Thus it was considered that the antipyretic activity of the free fatty acid may be due to the presence of arachidonic acid. Lipid-free earthworm powder was extracted with phosphate buffer (pH, 8.0, 0.1M) and all the proteins was salted out by ammonium sulfate. The crude precipitate was dialyzed and the impure proteins were eliminated at pH 5.4 and 4.6. The remaining protein solution was fractionated with various concentrations of acetone. The acetone fractions were identified by using S.D.S. polyacrylamide gel electrophoresis and disc gel electrophoresis. The precipitate at 85% acetone concentration and the remaining proteins in the supernatant did not exhibit the antipyretic activity.

• Journal of Microbiology and Biotechnology
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• 제22권9호
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• pp.1271-1278
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• 2012

Genetic fusion of two proteins frequently induces beneficial effects to the proteins, such as increased solubility, besides the combination of two protein functions. Here, we study the effects of the bacterial surface layer protein SgsE from Geobacillus stearothermophilus NRS 2004/3a on the folding of a C-terminally fused enhanced green fluorescent protein (EGFP) moiety. Although GFPs are generally unable to adopt a functional confirmation in the bacterial periplasm of Escherichia coli cells, we observed periplasmic fluorescence from a chimera of a 150-amino-acid N-terminal truncation of SgsE and EGFP. Based on this finding, unfolding and refolding kinetics of different S-layer-EGFP chimeras, a maltose binding protein-EGFP chimera, and sole EGFP were monitored using green fluorescence as indicator for the folded protein state. Calculated apparent rate constants for unfolding and refolding indicated different folding pathways for EGFP depending on the fusion partner used, and a clearly stabilizing effect was observed for the SgsE_C fusion moiety. Thermal stability, as determined by differential scanning calorimetry, and unfolding equilibria were found to be independent of the fused partner. We conclude that the stabilizing effect SgsE_C exerts on EGFP is due to a reduction of degrees of freedom for folding of EGFP in the fused state.

• Asian-Australasian Journal of Animal Sciences
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• 제20권3호
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• pp.307-312
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• 2007

The present study was carried out to investigate differentially expressed chicken muscle proteins using proteomics approach. More than 300 protein spots were investigated for the muscle samples in 2DE gels and the differentially expressed protein spots between pectoralis and peroneus longus muscles from Cornish and White Leghorn breeds were characterized by MALDI-TOF. In pectoralis muscles, PGAM1 protein was detected as differentially expressed between White Leghorn and Cornish breeds. On the other hand, 4 protein spots (SP22, nxf-2, SOD1, TNNI2) were differentially expressed between White Leghorn and Cornish breeds in peroneus longus muscles. These proteins assumed to be related with muscle development, growth, stress, and movements in chicken. In this experimental process, 2D reference map of the chicken muscle proteins was needed and 25 proteins, which were commonly expressed in both pectoralis and peroneus longus muscles in both breeds, were selected and characterized. Upon finishing the exact roles of the differentially expressed proteins, the identified 5 proteins will be used as valuable information for the fundamental mechanisms of muscle biology and underline genetics.

• 한국가축번식학회지
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• 제26권4호
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• pp.377-384
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• 2002

To investigate the expression patterns of proteins and growth factor signals in differentiated rabbit embryonic stem (ES) cells, ES cells with confluent stage grown of feeder layer and differentiated cells into embryoid bodies (EB) without feeder cell were applied to protein gel and Western blotting analysis. There were 66kDa and 28kDa specifically expressed in differentiated ES cell but not in undifferentiated ES cell while 25kDa protein band showed up in only undifferentiated ES cells. Also there were some difference of protein bands in several area of gel between differentiated and undifferentiated ES cells such as about 100 kDa, 50kDa and 27kDa areas, but there was no difference in band pattern of one-dimensional gel analysis between mouse ES cells and rabbit ES cells. IGF-I receptor and EGF receptor were expressed in differentiated cells and undifferentiated cells. And ICF-I and EGF were not expressed in both differentiated and undifferentiated cells. These results indicated that ES cells express their own proteins to inhibit differentiation while EB cells synthesize different proteins to differentiate, and 16F-I receptor and EGF receptor were expressed in both ES and EB cells probably for the different functions.

• 한국동물학회지
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• 제33권4호
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• pp.468-475
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• 1990

개선된 방법인 알칼리 가수분해와 얇은 층 cellulose전기 영동 (TLE)으로 in vitro 조건에서 쥐 뇌의 용해부분에서의 단백질 황산화의 특성을 추가로 조사하였다. 단백질 황산화는 방사능을 띤 [35 S] 3'-phosphoadenosine-5'-phosphosulfate (PAPS),Tris-maleate buffer (pH 8), MgCI$_2$, 그리고 쥐 뇌의 용해 단백질이 포함된 반응체 내에서 실행되었다. 황산화된 단백질들은 acetone에 의해 침전된 후 황산화된 아미노산을 얻기위해 알칼리 가수분해를 하였다. 더나아가 그 가수분해물을 TLE로 분리하고 황산화된 잔기들을 fluorography를 통해 확인하였다. 일차원 TLE의 fluorography에서는 tryosine-O-sulfate를 포함한 적어도 9개의 황산된 잔기들이 나타났다. tryosine-O-sulfate를 제외한 다른 잔기들은 아직 분명하게 밝혀지지 않고 있다. 이런 방법으로 단백질 황산기전달효소(PST)의 일반적 성질들 즉, PAPS농도, pH,반응온도,그리고 $Mg^2$+등의 효과를 재조사하였다. 이런 결고들은 쥐 뇌에서 각황산화된 잔기들에 해당하는 여러개의 PST의 존재가 가능하며 단백질 황산화가 tryosine뿐만 아니라 다른 잔기들에서도 일어날 수 있음을 암시한다.

• Asian-Australasian Journal of Animal Sciences
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• 제6권4호
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• pp.483-489
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• 1993

Bovine fibroblasts were cultured in Dulbecco's Modified Eagle's Medium and then treated with control, insulin (I, $1{\mu}g/ml$), cholera toxin (CT, 0.1-100 ng/ml) or CT (0.1-100 ng/ml) + I ($1{\mu}g/ml$). Cholera toxin, an activator of adenylate cyclase, significantly decreased insulin induced DNA synthesis (p<0.05). The modulation of DNA synthesis apparently involves events occurring in early stage of cell growth, at least between the first 4 and 8 hour of CT treatment. Insulin induced collagen as well as noncollagen synthesis in cell layer, however, these syntheses were reduced by addition of cholera toxin (p<0.05) but were not completely reduced. It is not clear whether the reduction of insulin-induced cell layer collagen or noncollagen proteins by CT is involved in the inhibitory effect on insulin-induced DNA synthesis. However, we could rule out the hypothesis that insulin-induced DNA synthesis is reduced by CT-induced cellular differentiation.

• Journal of Microbiology and Biotechnology
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• 제23권4호
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• pp.545-554
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• 2013

Plasmid isolation of kimchi-derived Weissella cibaria KLC140 revealed six different plasmids. The smallest plasmid, pKW2124, was DNA sequenced and characterized, showing 2,126 bp with a GC content of 36.39% and five putative open reading frames (ORFs). In silico analysis of these ORFs showed ORF1 encodes a putative replication protein similar to rolling circular replication proteins from other lactic acid bacteria. However, a single-stranded intermediate was not detected when S1 nuclease was treated, suggesting it may follow theta replication. Interestingly, the replication initiation site of this plasmid is 100% identical to other plasmids from lactic acid bacteria, suggesting it may function for replication initiation. To construct a surface layer expression vector, pTSLGFP, slpA encoding the surface layer protein from Lactobacillus acidophilus was PCR amplified and fused with the gfp gene, forming a SLGFP fused gene. The plasmid pKW2124 was cloned into the XbaI site of pUC19, forming an Weissella-E. coli shuttle vector pKUW22. NheI-linearized pTSLGFP was ligated into pKUWCAT containing pKUW22 and the chloramphenicol acetyltransferase gene from pEK104, resulting in an 8.6 kb pKWCSLGFP surface layer expression vector. After transformation of this vector into W. cibaria KLC140, a GFP fluorescence signal was detected on the surface of the transformant, substantiating production of SLGFP fused protein and its secretion. This is the first report for construction of a Weissella surface layer expression vector, which may be useful for surface layer production of beneficial proteins in Weissella.

• 대한기관식도과학회:학술대회논문집
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• 대한기관식도과학회 1993년도 제27차 학술대회 초록집
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• pp.88-88
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• 1993

현재 악성종양의 조직학적 진단에 면역조직화학반응의 검토가 많이 사용되는데 정상상피가 암으로의 진행시 점막상피세포의 주구성 단백성분에 있는 cytokeratin의 발현성에 변화가 일어난다고 알려져 있다. 이 cytokeratin은 현재 19종의 subclass가 알려져 있는데 각 subclass에 대응하는 항체를 이용하여 후두암세포의 기원을 알고 암조직의 분화도를 판정하고자 본 연구를 시행하였다. 연구방법은 정상성대조직 5예, 성대에 국한된 편평세포암 22예를 monoclonal anti-ck Ab와 polyclonal anti-ck Ab를 사용, Avidin-Biotin Peroxidase Complex method로 면역조직화학적 염색을 시행하였다. 그결과 정상후두상피에서 저분자량의 keratin은 표층을 제외한 심층부에서 양성반응을 보였다. 후두의 편평세포암은 상피성종양이며, 저분자량의 항ck 항체는 저분화암에서 양성반응을 보이고 고분자량의 항ck항체는 고분화암에서 양성반응을 보였다.

• Journal of the Korean Association of Oral and Maxillofacial Surgeons
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• 제28권6호
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• pp.421-433
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• 2002

In order to elucidate the pathogenesis of cleft lip and palate, first of all, it is necessary to understand the developmental mechanisms of growth factors and extracellular matrix proteins in the tissues of cleft lip and palate. We have performed immunohistochemical studies on human cleft lip and palate tissues to elucidate the pathogenetic implications of cleft lip and palate. 16 specimens from postnatal human cleft lip and palate subjects and 17 specimens from autopsy of prenatal human cleft lip and palate were fixed in 10% buffered formalin, embedded in paraffin. The sections were routinely stained by hematoxylin and eosin, also stained by PAS, and followed by immunohistochemical stainings using the antiseras of growth factors and extracellular matrix proteins such as PCNA, S-100, c-erb-B2, MMP-3, MMP-10, HSP-70, transglutaninase-C, E-cadherin, VEGF, vWF. Both the prenatal and postnatal specimens of cleft lip and palate showed dysplastic proliferation of the basal cell layer, increased infiltration of melanocytes into mucosal epithelium, sebaceous gland hyperplasia ingrowing into the muscular tissue of lip and palate, and fatty infiltration into the submucosal deep connective tissue. The strong reactions of MMP-3 and HSP-70 were detected in the tissues of cleft lip and palate, especially increased in degenerating muscle bundles, while the immunostainings of PCNA and c-erb-B2 were weakly positive in the tissues of cleft lip and palate. These data suggest that the retrogressive tissue degeneration around the cleft areas persistently exist during the prenatal and postnatal period after cleft formation, and the sebaceous gland hyperplasia and fatty infiltration with the intense expression of MMP-3 and HSP-70 is closely related to the muscular degeneration around the cleft area.

• Journal of Microbiology and Biotechnology
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• 제21권6호
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• pp.574-581
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• 2011

A Gram-positive bacterium was isolated from the saline soils of Jangpura (U.P.), India, and showed high-level of radiation-resistant property and survived upto 12.5 kGy dose of gamma radiation. The 16S rDNA sequence of this strain was examined, identified as Bacillus sp. strain HKG 112, and was submitted to the NCBI GenBank (Accession No. GQ925432). The mechanism of radiation resistance and gene level expression were examined by proteomic analysis of whole-cell extract. Two proteins, 38 kDa and 86.5 kDa excised from SDS-PAGE, which showed more significant changes after radiation exposure, were identified by MALDI-TOF as being flagellin and S-layer protein, respectively. Twenty selected 2-DE protein spots from the crude extracts of Bacillus sp. HKG 112, excised from 2- DE, were identified by liquid chromatography mass spectrometry (LC-MS) out of which 16 spots showed significant changes after radiation exposure and might be responsible for the radiation resistance property. Our results suggest that the different responses of some genes under radiation for the expression of radiation-dependent proteins could contribute to a physiological advantage and would be a significant initial step towards a fullsystem understanding of the radiation stress protection mechanisms of bacteria in different environments.