• IMMUNE NETWORK
• /
• 제4권3호
• /
• pp.184-189
• /
• 2004

Background: Hu syndrome, a neurological disorder, is characterized by the remote effect of small cell lung cancer on the neural degeneration. The suspicious effectors for this disease are anti-Hu autoantibodies or Hu-related CD8+ T lymphocytes. Interestingly, the same effectors have been suggested to act against tumor growth and this phenomenon may represent natural tumor immunity. For these diagnostic and therapeutic reasons, the demand for antibodies against Hu protein is rapidly growing. Methods: Polyclonal and monoclonal antibodies were generated using recombinant HuR protein. Western blot analyses were performed to check the specificity of generated antibodies using various recombinant proteins and cell lysates. Extracellular stimuli for HuR expression had been searched and HuR-associated proteins were isolated from polysome lysates and then separated in a 2-dimensional gel. Results: Polyclonal and monoclonal antibodies against HuR protein were generated and these antibodies showed HuR specificity. Antibodies were also useful to detect and immunoprecipitate endogenous HuR protein in Jurkat and BJAB. This report also revealed that TNF-${\alpha}$ treatment in BJAB up-regulated HuR expression. Lastly, protein profile in HuR-associated mRNAprotein complexes was mapped by 2-dimensional gel electrophoresis. Conclusion: This study reported that new antibodies against HuR protein were successfully generated. Currently, project to develop a diagnostic kit is in process. Also, this report showed that TNF-${\alpha}$ up-regulated HuR expression in BJAB and protein profile associated with HuR protein was mapped.

• Journal of Microbiology and Biotechnology
• /
• 제24권12호
• /
• pp.1719-1727
• /
• 2014

In the present study, whispovirus immediate early 1 promoter (ie-1) was used to initiate surface expression of the hemagglutinin (HA) protein of Egyptian H5N1 avian influenza virus (AIV) by using the baculovirus expression vector system. The HA gene and whispovirus ie-1 promoter sequence were synthesized as a fused expression cassette (ie1-HA) and successfully cloned into the pFastBac-1 transfer vector. The recombinant vector was transformed into DH10Bac competent cells, and the recombinant bacmid was generated via site-specific transposition. The recombinant bacmid was used for transfection of Spodoptera frugiperda (Sf-9) insect cells to construct the recombinant baculovirus and to induce expression of the HA protein of H5N1 AIV. The recombinant glycoprotein expressed in Sf-9 cells showed hemadsorption activity. Hemagglutination activity was also detected in both extra- and intracellular recombinant HAs. Both the HA and hemadsorption activities were inhibited by reference polyclonal anti-H5 sera. Significant expression of the recombinant protein was observed on the surface of infected insect cells by using immunofluorescence. SDS-PAGE analysis of the expressed protein revealed the presence of a visually distinguishable band of ~63 kDa in size, which was absent in the non-infected cell control. Western blot analysis confirmed that the distinct 63 kDa band corresponded to the recombinant HA glycoprotein of H5N1 AIV. This study reports the successful expression of the HA protein of H5N1 AIV. The expressed protein was displayed on the plasma membrane of infected insect cells under the control of whispovirus ie-1 promoter by using the baculovirus expression vector system.

• Asian Pacific Journal of Cancer Prevention
• /
• 제13권12호
• /
• pp.6239-6244
• /
• 2012

Background: The basal cell carcinoma (BCC) is the most common non-melanoma skin cancer (NMSK). BCC might develop because of the faulty cell cycle arrest. $p15^{INK4b}$ is a tumor suppressor gene, involved in cell cycle arrest and inactivated in most human cancers. The role of $p15^{INK4b}$ protein expression in the genesis of BCC is as yet unknown. In a previous study we showed the absence of $p15^{INK4b}$ expression in the majority of tissue microarray cores of cutaneous squamous cell carcinoma (SCCs), another type of non-melanoma skin cancer, indicating that $p15^{INK4b}$ could possibly be involved in the pathogenesis of cutaneous SCC. The aim of this study was to investigate $p15^{INK4b}$ protein expression in BCCs. Materials and Method: Protein expression of $p15^{INK4b}$ in 35 cases of BCC tissue arrays and 19 cases of normal human skin tissue was studied using an immunohistochemical approach. Results: The expression of $p15^{INK4b}$ was not significantly different in the BCC cases as compared with normal human skin (p=0.356; p>0.05). In addition, there were no significant relationship between clinicopathologic variables of patients (age and sex) and $p15^{INK4b}$ protein expression. Conclusions: Our finding may indicate that $p15^{INK4b}$ protein expression does not play a role in the genesis of BCC.

• Asian Pacific Journal of Cancer Prevention
• /
• 제15권20호
• /
• pp.8861-8869
• /
• 2014

Background: The prognostic value of Bcl-2 protein expression in non-small cell lung cancer (NSCLC) is under debate. We therefore systematically reviewed the evidence for Bcl-2 protein effects on NSCLC survival to elucidate this issue. Materials and Methods: An electronic search in Pubmed and Embase complemented by manual searches in article references were conducted to identify eligible studies to evaluate the association between Bcl-2 protein expression and overall survival (OS) as well as disease free survival (DFS) of NSCLC patients. Combined hazard ratios (HRs) with corresponding 95% confidence intervals (95%CIs) were pooled using the random-effects model. Results: A total of 50 trials (including 52 cohorts) encompassing 7,765 patients were pooled in the meta-analysis regarding Bcl-2 expression and OS of NSCLC patients. High expression of Bcl-2 protein had a favorable impact (HR=0.76, 95%CI=0.67-0.86). In the group of Bcl-2 expression and DFS, 11 studies including 2,634 patients were included. The synthesized result indicated high expression of Bcl-2 protein might predict good DFS (HR=0.85, 95%CI=0.75-0.95). Conclusions: Our present meta-analysis demonstrated favorable prognostic values of Bcl-2 expression in patients with NSCLC. Further prospective trails are welcomed to validate the utility of assessing Bcl-2 in NSCLC patient management.

• Journal of Gastric Cancer
• /
• 제3권2호
• /
• pp.75-79
• /
• 2003

Purpose: Evidence exists that dysregulation of apoptosis is involved in the pathogenesis of cancer development. The Bcl-$x_{L}$/Bcl-2-associated death promoter (BAD), a member of the Bcl-2 family, is a critical regulatory component of the intrinsic cell-death pathway that exerts its pro-apoptotic effect upon heterodimerization with anti-apoptotic proteins Bcl-2 and Bcl-$X_{L}$. Expression of the BAD protein has been reported in several cancer types, but not in stomach cancer. The aim of this study was to explore the expression status of the BAD protein in gastric carcinomas. Materials and Methods: In the current study, we analyzed the expression of the BAD protein in 60 advanced gastric adenocarcinomas by using immunohistochemistry and a tissue microarray approach. Results: Immunopositivity (defined as $\geq\30\%$) was observed for the BAD protein in 57 ($95\%$) of the 60 cancers. Normal gastric mucosal cells showed weaker expressions of the BAD protein than gastric carcinomas. Conclusion: Taken together, these results suggest that stomach cancer cells in vivo may need BAD protein expression for apoptosis. Also, the higher expression of the BAD protein in stomach cancer cells than in normal gastric mucosal cells suggests that apoptosis might be easily triggered in susceptible stomach cancer cells, thereby producing selective pressure to make more apoptosis-resistant cells during tumor development.

• International Journal of Industrial Entomology and Biomaterials
• /
• 제22권2호
• /
• pp.59-64
• /
• 2011

Protein kinase C (PKC) is involved in many cellular signaling pathways, it participates in many physiological processes, such as cell cycle, growth, proliferation, differentiation and apoptosis. To investigate the effect of PKC on the silkworm midgut tissue infection of Bombyx mori parvo-like virus (BmPLV), a B. mori atypical protein kinase C (BmaPKC) gene was cloned from larval midgut tissue, expressed in E. coli and purified. Additionally, the BmPLV susceptible silkworm strain and resistant silkworm strain were used to test the effect of the B. mori infection on BmPLV. The result showed that BmaPKC encodes a predicted 586 amino acid protein, which contains a C-terminal kinase domain and an N-terminal regulatory domain. The maximum expression amount of the soluble (His)6-tagged fusion protein was detected after 0.8 mmol/L IPTG was added and cultured at $21^{\circ}C$. The (His) 6-tagged fusion protein revealed about 73 kDa molecular weight which confirmed by western blot and mass spectrography. Furthermore BmaPKC protein were detected at 0-72 h post-infection in BmPLVinfected larval midgut tissue, western blot showed that as time went on, the expression of BmaPKC increased gradually in susceptible strain, the expression quantity on 72 h is 5 times of 0 h. However, in resistant strain, the expression quantity is slightly lower than susceptible strain. But no significant change in resistant strain was observed as time went on. The available data suggest that BmaPKC may involve in the regulation of BmPLV proliferation.

• KSBB Journal
• /
• 제24권1호
• /
• pp.96-100
• /
• 2009

본 연구에서는 저온에서의 재조합 단백질 생산성 향상을 위하여 저온에서 RNA 샤페론 활성을 지닌다고 알려진 CspA 단백질의 발현이 서로 다른 온도에서 대장균의 성장 및 GFP의 발현 속도에 어떻게 영향을 미치는지를 살펴보았다. $20^{\circ}C$, $25^{\circ}C$, $37^{\circ}C$에서는 세포 성장 및 GFP의 생산이 CspA의 발현에 영향을 받지 않았으나, $15^{\circ}C$에서는 GFP의 총 생산성이 CspA의 동시 발현에 의해 향상되었으며 이는 세포 성장 속도의 향상에 기인함을 확인하였다. 결론적으로 CspA의 발현은 $15^{\circ}C$에서 세포 당 재조합 단백질 생산량의 증가에는 영향을 미치지 않으나, 즉 재조합 단백질의 번역 효율에는 큰 영향을 미치지 않으나, 대장균 성장 속도에 영향을 미치며, 이를 통해 재조합 단백질의 총 생산량 향상을 유도 할 수 있을 것으로 기대된다.

• Biotechnology and Bioprocess Engineering:BBE
• /
• 제9권3호
• /
• pp.184-190
• /
• 2004

The influences of impeller types on morphology and protein expression were investigated in a submerged culture of Aspergillus oryzae. The impeller types strongly affected mycelial morphology and protein production in batch and fed-batch fermentations. Cells that were cultured by propeller agitation grew in the form of a pellet, whereas cells that were cultured by turbine agitation grew in a freely dispersed-hyphal manner and in a clumped form. Pellet-grown cells showed high levels of protein production for both the intracellularly heterologous protein (${\beta}$-glucuronidase) and the extracellularly homologous protein (${\alpha}$-amylase). The feeding mode of the carbon source also influenced the morphological distribution and protein expression in fed-batch fermentation of A. oryzae. Pulsed-feeding mainly showed high protein expression and homogeneous distribution of pellet whereas continuous feeding resulted in less protein expression and heterogeneous distribution with pellet and dispersed-hyphae. The pellet growth with propeller agitation paralleling with the pulsed-feeding of carbon source showed a high level of protein production in the submerged fed-batch fermentation of recombinant A. oryzae.

• 한국작물학회지
• /
• 제60권1호
• /
• pp.41-46
• /
• 2015

본 연구는 국내 육성 품종인 태광콩의 등숙기에 따른 단백질 발현 양상을 비교함으로써 등숙기 단백질 발현의 차이에 대한 기초자료를 얻고자 수행하였다. 동한 개화 후 종실의 등숙이 진행됨에 따라서 단백질 발현 양상이 세가지 경향으로 나뉘어 지는 것을 확인하였다. 첫 번째는 등숙이 진행됨에 따라서 단백질 발현 정도가 증가하다가 감소되며, 두 번째는 증가와 감소의 시기가 성숙기에 이루어지며, 세번째는 등숙기부터 성숙기까지 점진적으로 증가하는 것이다. 이러한 현상은 단백질의 기능에 따라 달라지는 것으로 사료된다. 등숙 초기에는 등숙에 필요한 단백질의 발현이 증가할 것이며 등숙 후기에는 저장단백질의 발현이 증가할 것으로 사료된다. 따라서 향후 좀 더 많은 수의 단백질 spot 들을 동정하여 어떤 기능을 가진 단백질이 등숙기에 따라 단백질의 발현 양상이 달라지는지는 좀 더 면밀히 관찰할 필요성이 있다고 사료된다.

• 한국식품영양학회지
• /
• 제12권2호
• /
• pp.184-190
• /
• 1999

Soybean protein consists of two major components $\beta$-conglycinin and glycinin which together consti-tute 70% of the total seed storage protein at maturity. $\beta$-Conglycinin is trimeric glycoprotein and for-med by the assembly of various combinations of three subunits $\alpha$,$\alpha$' and $\beta$ which have molecular weig-hts of 69,000, 72,000 and 42,000, respectively. Recently $\beta$-conglycinin was identified as powerful LDL lip-oprotein receptor activation hypercholesterolemia and major allergenic proteins. To investigate these reasons we constructed an expression system of cDNA encoding $\alpha$-subunit of $\beta$-conglycinin in Escherichia coli and purified the expressed protein. The pro-$\beta$-conglycinin synthesized in Escherichia coli BL 21 (DE3)comprised approximately 15% of the total bacterial proteins and the expressed protein are formed sol-uble and trimer such as native protein in Escherichia coli cells. The highly expressed protein was purified to homogeneity by salt precipitation with 20~40 % ammonium sulfate ion-exchange chromatography with Q-sepharose and hydrophobic column chromatography with Butyltoyopearl.