• Journal of Microbiology and Biotechnology
• /
• 제29권5호
• /
• pp.813-819
• /
• 2019

Middle East respiratory syndrome coronavirus (MERS-CoV) induces severe respiratory impairment with a reported mortality rate of ~36% in humans. The absence of clinically available MERS-CoV vaccines and treatments to date has resulted in uncontrolled incidence and propagation of the virus. In vaccine design, fusion with the IgG Fc domain is reported to increase the immunogenicity of various vaccine antigens. However, limited reports have documented the potential negative effects of Fc fusion on vaccine antigens. To determine whether Fc fusion affects the immunogenicity of MERS-CoV antigen, we constructed a Fcassociated MERS-CoV spike protein (eS770-Fc, 110 kDa), whereby human IgG4 Fc domain was fused to MERS-CoV spike protein (eS770) via a Gly/Pro linker using baculovirus as the expression system. For comparative analyses, two eS770 proteins lacking the IgG4 Fc domain were generated using the IdeS protease ($eS770-{\Delta}Fc$) or His tag attachment (eS770-His) and the immunogenicity of the above constructs were examined following intramuscular immunization in mice. Contrary to expectations, non-Fc spike proteins ($eS770-{\Delta}Fc$, eS770-His; 90 kDa) showed higher immunogenicity than the Fc fusion protein (eS770-Fc). Moreover, unlike non-Fc spike proteins, eS770-Fc immunization did not elicit neutralizing antibodies against MERS-CoV. The lower immunogenicity of Fc-fused eS770 was related to alterations in the structural conformation of the spike protein. Taken together, our results indicate that IgG Fc fusion reduces the immunogenicity of eS770 by interfering with the proper folding structure.

• 한국동물위생학회지
• /
• 제47권1호
• /
• pp.9-17
• /
• 2024

Porcine epidemic diarrhea (PED) is a highly contagious enteric viral disease of pigs with watery diarrhea in piglets, which ultimately results in huge economic losses in the swine industry. The spike (S) protein plays an important role in viral pathogenicity, tissue tropism, infection, dissemination and the trypsin-dependent proliferation of the PED virus (PEDV). In the present study, we determined the full-length spike (S) gene sequences of twenty PEDV field strains detected in Jeonbuk province in 2022. Phylogenetic analysis showed that the twenty PEDV field strains were classified into G2b group and shared 98.6~100% of nucleotide homology and 97.4~100% of amino acid homology with each other. Mutations of amino acid sequences on the neutralizing epitope of S protein were observed in the twenty field strains compared to the previous vaccine strain SM-98-1 (G1a group). Therefore, these amino acid mutations in the PEDV S protein may result in a new genotype of the virus and highly pathogenic virus, so continuous monitoring is required.

• 대한수의학회지
• /
• 제36권3호
• /
• pp.627-633
• /
• 1996

To analyze the genomic diversity of transmissible gastroenteritis virus (TGEV), the N-terminal half of the spike (S) glycoprotein gene and nonstructural protein gene (open reading frames 3 and 3-1) were amplified by reverse transcriptase reaction and polymerase chain reaction (RT-PCR), and analyzed using restriction fragment length polymorphism (RFLP) patterns of the amplified DNA. In this study, TGEV Miller (M6) and Purdue (P115) strains were used as reference strains, and two vaccine strains (MSV and STC3) and four Korea isolates (P44, VRI-WP, VRI-41, and VRI-48) were analyzed. All TGEV strains were amplified with three TGEV primer pairs. Although there was some exception in RFLP analysis, this method differentiated TGEV strains into following groups : Miller group (M6 and MSV), Purdue group (PUS, STC3, P44, VRI-WP, VRI-41, and VRI-48). Using Sau3AI and SspI, VRI-48 was differentiated from the Miller and Purdue type viruses. The RT/PCR in conjuction with RFLP analysis was a rapid and valuable tool for differentiating several strains of TGEV. This study revealed the occurences of distinct difference in genome of TGEV strains.

• Genomics & Informatics
• /
• 제18권4호
• /
• pp.44.1-44.7
• /
• 2020

The severity of coronavirus disease 2019 (COVID-19), caused by the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), greatly varies from patient to patient. In the present study, we explored and compared mutation profiles of SARS-CoV-2 isolated from mildly affected and severely affected COVID-19 patients in order to explore any relationship between mutation profile and disease severity. Genomic sequences of SARS-CoV-2 were downloaded from Global Initiative on Sharing Avian Influenza Data (GISAID) database. With the help of Genome Detective Coronavirus Typing Tool, genomic sequences were aligned with the Wuhan seafood market pneumonia virus reference sequence and all the mutations were identified. Distribution of mutant variants was then compared between mildly and severely affected groups. Among the numerous mutations detected, 14408C>T and 23403A>G mutations resulting in RNA-dependent RNA polymerase (RdRp) P323L and spike protein D614G mutations, respectively, were found predominantly in severely affected group (>82%) compared with mildly affected group (<46%, p < 0.001). The 241C>T mutation in the non-coding region of the genome was also found predominantly in severely affected group (p < 0.001). The 3037C>T, a silent mutation, also appeared in relatively high frequency in severely affected group compared with mildly affected group, but the difference was not statistically significant (p = 0.06). We concluded that spike protein D614G and RdRp P323L mutations in SARS-CoV-2 are associated with severity of COVID-19. Further studies will be required to explore whether these mutations have any impact on the severity of disease.

• ALGAE
• /
• 제38권4호
• /
• pp.295-306
• /
• 2023

Human coronavirus diseases, particularly severe acute respiratory syndrome coronavirus 2, still remain a persistent public health issue, and many recent studies are focusing on the quest for new leads against coronaviruses. To contribute to this growing pool of knowledge and explore the available marine natural products against coronaviruses, this study investigated the antiviral effects of fucoxanthin isolated from Sargassum siliquastrum-a brown alga found on Jeju Island, South Korea. The antiviral effects of fucoxanthin were confirmed in severe acute respiratory syndrome coronavirus 2-infected Vero cells, and its structural characteristics were verified in silico using molecular docking and molecular dynamic simulations and in vitro colorimetric method. Fucoxanthin inhibited the infection in a concentration-dependent manner, without showing cytotoxicity. Molecular docking simulations revealed that fucoxanthin binds to the angiotensinconverting enzyme 2-spike protein (binding energy -318.306 kcal mol^-1) and main protease (binding energy -205.118 kcal mol^-1). Moreover, molecular dynamic simulations showed that fucoxanthin remains docked to angiotensin-converting enzyme 2-spike protein for 20 ns, whereas it breaks away from main protease after 3 ns. Also, the in silico prediction of the fucoxanthin was verified through the in vitro colorimetric method by inhibiting the binding between angiotensinconverting enzyme 2 and spike protein in a concentration-dependent manner. These results indicate that fucoxanthin exhibits antiviral effects against severe acute respiratory syndrome coronavirus 2 by blocking the entry of the virus. Therefore, fucoxanthin from S. siliquastrum can be a potential candidate for treating coronavirus infection.

• BMB Reports
• /
• 제52권6호
• /
• pp.397-402
• /
• 2019

Middle East respiratory syndrome coronavirus (MERS-CoV) uses the spike (S) glycoprotein to recognize and enter target cells. In this study, we selected two epitope peptide sequences within the receptor binding domain (RBD) of the MERS-CoV S protein. We used a complex consisting of the epitope peptide of the MERS-CoV S protein and CpG-DNA encapsulated in liposome complex to immunize mice, and produced the monoclonal antibodies 506-2G10G5 and 492-1G10E4E2. The western blotting data showed that both monoclonal antibodies detected the S protein and immunoprecipitated the native form of the S protein. Indirect immunofluorescence and confocal analysis suggested strong reactivity of the antibodies towards the S protein of MERS-CoV virus infected Vero cells. Furthermore, the 506-2G10G5 monoclonal antibody significantly reduced plaque formation in MERS-CoV infected Vero cells compared to normal mouse IgG and 492-1G10E4E2. Thus, we successfully produced a monoclonal antibody directed against the RBD domain of the S protein which could be used in the development of diagnostics and therapeutic applications in the future.

• 한국미생물·생명공학회지
• /
• 제39권2호
• /
• pp.140-145
• /
• 2011

돼지유행성설사 바이러스(porcine epidemic diarrhea virus: PEDV)는 자돈에게 감염 시 수양성설사를 동반한 급성 장염을 유발하며 매우 높은 폐사율을 보이는 그룹 1 코로나바이러스이다. PEDV는 다른 그룹 1 코로나바이러스와 마찬가지로 숙주 세포에 감염 시 aminopeptidase N (APN)을 세포 수용체로 이용한다고 알려져 있다. 코로나바이러스의 spike(S) 단백질은 숙주세포의 표면에 부착과 관련하여 감염 개시에 있어 중요한 역할을 하는 것으로 알려져 있으며 특히 S 단백질의 S1 도메인은 세포 수용체에 특이적인 결합을 매개하는 수용체 결합 도메인(receptor binding domain: RBD)을 포함하고 있는 것으로 알려져 있다. 이미 많은 코로나바이러스의 RBD의 위치가 확인되어져 있지만 PEDV의 RBD에 대해서는 아직까지 알려진 바가 없다. 본 연구에서는 돼지 APN 수용체와 결합을 매개하는 PEDV의 RBD를 규명하기 위해 S1 도메인을 주형으로 하는 일련의 재조합 truncated variant들을 제작하였고 각각의 truncated들이 실제로 pAPN과의 결합을 이루는지에 대하여 실험을 통해 확인하였다. 그 결과 S1 도메인의 N 말단 부분이 pAPN과의 결합에서 중요한 부위임을 확인할 수 있었다. 본 연구에서 도출된 결과는 향후 PEDV의 S 단백질과 pAPN간의 분자적 상호작용을 이해하는 데에 도움을 줄 것으로 판단된다.

• 한국작물학회지
• /
• 제58권3호
• /
• pp.239-252
• /
• 2013

국산밀을 재배하는 전국 175농가를 대상으로 2년 동안 농업형질과 원맥 특성을 조사 한 결과, 간장, 수장, 경수와 이형주율은 년차간 차이를 나타내었다. 시비량이 많았던 남부지역에서 재배된 밀의 간장이 중북부 지역 보다 길었으며, 추비시기는 간장에 영향이 없었지만, 3월 중순 이후에 늦게 추비를 준 농가의 수장은 다른 농가에 비하여 길었으나 경수는 감소하였다. 추비 횟수는 농업형질에 영향을 미치지 않는 것으로 나타났다. 농업형질과 마찬가지로 원맥 특성도 연차간 변이가 있었으며, 남부지역에서 재배된 밀은 중북부지역에서 재배된 밀에 비하여 천립중이 높았으며, 리터중은 낮았으며, 전라북도는 다른 지역에 비하여 낮은 회분 함량과 높은 단백질 함량을 나타냈으며, 전라남도는 다른 지역에 비하여 높은 회분 함량을 나타내었다. 추비시기가 리터중에는 영향을 주었지만 천립중과 회분 및 단백질 함량에는 영향을 주지 않았으며, 추비 횟수도 원맥 특성에 영향이 없는 것으로 나타났다. 시비량에 따라 간장이 증가하고, 천립중과 단백질 함량이 증가하였으며, 수장과 회분함량은 시비량과 부의 상관을 나타내었다. 단백질 함량은 천립중과 정의 상관을 나타내었지만 회분 함량은 천립중 및 리터중과 부의 상관을 나타내었다. 간장이 증가할수록 천립중은 증가하고 리터중은 감소하였으며, 수장은 천립중과 부의 상관을 나타내었다. 시비량이 간장 및 원맥 특성에 영향을 미치기 때문에 품종별 용도에 맞는 밀 품종을 재배하기 위해서는 적정 시비기술에 대한 농가 지도가 필요하다. 또한, 국산 밀의 생산성 증진과 품질 향상 및 균일성 확보를 위해서는 용도별 가공적성에 적합한 품종에 대한 집단 재배단지를 조성하고 철저한 생산 관리와 체계적인 수확 후 관리 구축에 대한 종합적인 고려가 있어야 할 것이다.

• IMMUNE NETWORK
• /
• 제21권2호
• /
• pp.16.1-16.8
• /
• 2021

Patients with severe coronavirus disease 2019 (COVID-19) demonstrate dysregulated immune responses including exacerbated neutrophil functions. Massive neutrophil infiltrations accompanying neutrophil extracellular trap (NET) formations are also observed in patients with severe COVID-19. However, the mechanism underlying severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2)-induced NET formation has not yet been elucidated. Here we show that 2 viral proteins encoded by SARS-CoV-2, the nucleocapsid protein and the whole spike protein, induce NET formation from neutrophils. NET formation was ROSindependent and was completely inhibited by the spleen tyrosine kinase inhibition. The inhibition of p38 MAPK, protein kinase C, and JNK signaling pathways also inhibited viral protein-induced NET formation. Our findings demonstrate one method by which SARSCoV-2 evades innate immunity and provide a potential target for therapeutics to treat patients with severe COVID-19.

• BMB Reports
• /
• 제41권3호
• /
• pp.217-222
• /
• 2008

Based on a five-letter model of the 20 amino acids, we propose a new 2-D graphical representation of protein sequence. Then we transform the 2-D graphical representation into a numerical characterization that will facilitate quantitative comparisons of protein sequences. As an application, we construct the phylogenetic tree of 56 coronavirus spike proteins. The resulting tree agrees well with the established taxonomic groups.