Severe acute respiratory syndrome (SARS) is a severe respiratory infectious disease caused by a novel human coronavirus, SARS-CoV. The 3CL protease is a key enzyme in the proteolytic processing of replicase polyprotein precursors, pp1a and pp1ab, which mediate all the functions required for viral genomic replication and transcription. Therefore, this enzyme is a target for the development of chemotherapeutic agents against SARS. A large quantity of active SARS-3CL protease is required for development of anti-SARS agents. Here we have constructed overexpression vector for the production of the SARS-3CL protease. The gene encoding SARS-3CL protease was amplified using polymerase chain reaction and cloned into the pET29a expression vector, resulting in pET29a/SARS-3CLP. Recombinant SARS-3CL protease was successfully synthesized by the dialysis mode of the cell-free protein expression system, and purified by three-step fast protein liquid chromatography using HighQ and MonoP column chromatographies and Sephacryl S-300 gel filtration. In addition, the produced SARS-3CL protease was found to be an active mature form. This study provides efficient methods not only for the development of anti-SARS materials from natural sources, but also for the study of basic properties of the SARS-3CL protease.
Kim, Jong-Sik;Kang, Na-Kyung;Park, Seon-Mi;Lee, Eun-Joo;Chung, Kyung Tae
Journal of Life Science
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v.30
no.8
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pp.731-741
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2020
Coronavirus disease 19 (COVID-19) is caused by SARS-CoV-2 (Severe Acute Respiratory SyndromeCoronavirus 2). To date, seven coronaviruses that can infect humans were reported. Among them, infections with four coronavirus strains (HCoV-229E, HCoV-OC43, HCoV-NL63, and HCoV-HKU1) resulted in mild symptoms such as common cold, whereas SARS-CoV and MERS-CoV caused severe symptoms and epidemics in 2002 and 2012, respectively. In the most recent, SARS-CoV-2 was first reported in Wuhan, China in December 2019 and became a notorious cause of the ongoing global pandemics. To diagnose, treat, and prevent COVID-19, the development of rapid and accurate diagnostic tools, specific therapeutic drugs, and safe vaccines essentially are required. In order to develop these powerful tools, it is prerequisite to understand a phenotype, a genotype, and life cycle of SARS-CoV-2. Diagnostic techniques have been developing rapidly around world and many countries take the fast track system to accelerate approval. Approved diagnostic devices are rapidly growing facing to urgent demand to identify carriers. Currently developed commercial diagnostic devices are divided into mainly two categories: molecular assay and serological & immunological assay. Molecular assays begins the reverse transcription step following polymerase chain reaction or isothermal amplification. Immunological assay targets SARS-CoV-2 antigen or anti-SARS-CoV-2 antibody of samples. In this review, we summarize the phenotype, genome structure and gene expression of SARS-CoV-2 and provide the knowledge on various diagnostic techniques for SARS-CoV-2.
Kim Cho-Ja;Yoo Hye-Ra;Yoo Myung-Sook;Kwon Bo-Eun;Hwang Kyung-Ja
Journal of Korean Academy of Nursing
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v.36
no.4
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pp.596-603
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2006
Purpose. This study examined Korean clinical nurses' intentions to care for SARS patients and identify determinants of the intentions. Theory of planned behavior was the framework to explain the intentions of Korean nurses for SARS patients care. Methods. A convenient sample of six hundreds and seventy nine clinical nurses from four university-affiliated hospitals located in Seoul and in Kyung-gi province was used. Self-administered (83-items) questionnaire was used to collect data. Intentions, attitude, subjective norm, perceive behavioral control, behavioral beliefs, normative beliefs, and control beliefs were the study variables. All items were measured using 7point Likert scale (-3 to +3). Data were analyzed using descriptive statistics, Pearson correlation method, and stepwise multiple regression methods. Results. Intentions and attitudes toward SARS patient care among Korean clinical nurses were moderate, but their subjective norm and perceive behavioral control of SARS patients care were negative. Stepwise multiple regression analysis indicated that attitude toward SARS patient care, perceived behavioral control, subjective norm were the determinants of the intentions for SARS patients care as theory proposed. Among the behavioral beliefs, 'SARS-patient caring would be a new experience', 'during SARS-patient caring, I should be apart from my family', 'after completing SARS-patient caring, I would be proud of myself being able to cope with a stressful event' and 'with my SARS-patient caring, patients could recover from SARS' were the significant determinants. Among the normative beliefs, colleague approval, spouse approval, and physician approval were significant determinants of the intentions. Among the control beliefs, 'SARS-patient caring would be a challenge' 'SARS-patient caring is a professional responsibility', 'tension during the care of SARS patients' and 'support from team members' were the significant determinants of the intentions. Conclusions. Korean clinical nurses in this study were not willing to care for SARS patients and showed negative attitude toward the care. They believed their friends and family were not approved their care for SARS patients. Nurses were in conflicts between professional responsibilities to care for SARS patients and personal safety. This study was the first to understand stress and burden of Korean clinical nurses who are in front line to care for newly developed communicable disease such as SARS. Under the circumstance where several fatal communicable diseases are predictable, conflicts between professional responsibility and their personal risks should be taken into considerations by nurses themselves and by nursing administrators in order to improve quality of care.
Background : In Korea, the effort to control the spread of severe acute respiratory syndrome (SARS) has succeeded, but we have to maintain vigilance against SARS and other infectious diseases, because many experts warn that SARS may return or that other infectious agents could spread globally. The purpose of this study is to describe the degree of knowledge of hospital employees, assuming full responsibility for treating and attending to SARS suspected case, and to elucidate the attitude to SARS using questionnaires. Method : All employees of Kangwon university hospital were eligible for this study. We used validated questionnaires to survey demographic characteristics, the degree of knowledge of SARS, intention of treating and attending suspected cases, influence and effects of SARS on the community, and acquisition of information about SARS. Result : Based on the 251 completed surveys, out of 280 returned the score for knowledge about SARS was $72.0{\pm}13.7$, 50.6% responded positively about joining a SARS team, and 23.1% were opposed to the designation for quarantining. There was no relationship between the degree of knowledge of SARS and the attitudes to it. Most employees wanted to know how to manage SARS patients and how to prevent SARS. Conclusion : The attitudes to SARS were not affected by the level of knowledge of SARS. We suggest that national policies about SARS and other infectious diseases should be promulgated using the mass media and internet to provide information on rapidly spreading infectious agents, especially about the proper protocol for treating suspected cases and the appropriate steps to prevent or reduce the risk.
Objectives: Control methods against severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) aerosols have been introduced. Airborne spreading theories for SARS-CoV-2 were analyzed in this study. Methods: Control methods for airborne microorganisms were discussed. Studies on theoretical estimations for airborne spreading of SARS-CoV-2 were presented and analyzed. Analytic calculations were conducted for explaining control techniques for airborne microorganisms. Results: Control methods for SARS-CoV-2 aerosols can include physical or biological procedures. Characterization of SARS-CoV-2 aerosols and massive clustering infection cases of COVID-19 support the airborne spreading theories of SARS-CoV-2. It is necessary to consider the disadvantages of control methods for airborne microorganisms. Conclusions: A study on control methods against bioaerosols is necessary to prevent the spreading of viruses. Airborne spreading theories of SARS-CoV-2 were supported by the current evidence, but further studies are needed to confirm these theories.
The sudden appearance and potential lethality of severe acute respiratory syndrome (SARS)-associated coronavirus (SARS-CoV) in humans has resulted in a focusing of new attention on the determination of both its origins and evolution. The relationship existing between SARS-CoV and other groups of coronaviruses was determined via analyses of phylogenetic trees and comparative genomic analyses of the coronavirus genes: polymerase (Orflab), spike (S), envelope (E), membrane (M) and nucleocapsid (N). Although the coronaviruses are traditionally classed into 3 groups, with SARS-CoV forming a $4^{th}$ group, the phylogenetic position and origins of SARS-CoV remain a matter of some controversy. Thus, we conducted extensive phylogeneitc analyses of the genes common to all coronavirus groups, using the Neighbor-joining, Maximum-likelihood, and Bayesian methods. Our data evidenced largely identical topology for all of the obtained phylogenetic trees, thus supporting the hypothesis that the relationship existing between SARS-CoV and group 2 coronavirus is a monophyletic one. Additional comparative genomic studies, including sequence similarity and protein secondary structure analyses, suggested that SARS-Co V may bear a closer relationship with group 2 than with the other coronavirus groups. Although our data strongly suggest that group 2 coronaviruses are most closely related with SARS-CoV, further and more detailed analyses may provide us with an increased amount of information regarding the origins and evolution of the coronaviruses, most notably SARS-CoV.
Jaber, Abdullah All;Chowdhury, Zeshan Mahmud;Bhattacharjee, Arittra;Mourin, Muntahi;Keya, Chaman Ara;Bhuyan, Zaied Ahmed
Genomics & Informatics
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v.19
no.4
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pp.48.1-48.10
/
2021
Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) encodes small envelope protein (E) that plays a major role in viral assembly, release, pathogenesis, and host inflammation. Previous studies demonstrated that pyrazine ring containing amiloride analogs inhibit this protein in different types of coronavirus including SARS-CoV-1 small envelope protein E (SARS-CoV-1 E). SARS-CoV-1 E has 93.42% sequence identity with SARS-CoV-2 E and shared a conserved domain NS3/small envelope protein (NS3_envE). Amiloride analog hexamethylene amiloride (HMA) can inhibit SARS-CoV-1 E. Therefore, we performed molecular docking and dynamics simulations to explore whether amiloride analogs are effective in inhibiting SARS-CoV-2 E. To do so, SARS-CoV-1 E and SARS-CoV-2 E proteins were taken as receptors while HMA and 3-amino-5-(azepan-1-yl)-N-(diaminomethylidene)-6-pyrimidin-5-ylpyrazine-2-carboxamide (3A5NP2C) were selected as ligands. Molecular docking simulation showed higher binding affinity scores of HMA and 3A5NP2C for SARS-CoV-2 E than SARS-CoV-1 E. Moreover, HMA and 3A5NP2C engaged more amino acids in SARS-CoV-2 E. Molecular dynamics simulation for 1 ㎲ (1,000 ns) revealed that these ligands could alter the native structure of the proteins and their flexibility. Our study suggests that suitable amiloride analogs might yield a prospective drug against coronavirus disease 2019.
Na, Eun-Jee;Lee, Sook-Young;Kim, Hak Jun;Oem, Jae-Ku
Journal of Veterinary Science
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v.22
no.1
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pp.12.1-12.11
/
2021
Background: Bats have been considered natural reservoirs for several pathogenic human coronaviruses (CoVs) in the last two decades. Recently, a bat CoV was detected in the Republic of Korea; its entire genome was sequenced and reported to be genetically similar to that of the severe acute respiratory syndrome CoV (SARS-CoV). Objectives: The objective of this study was to compare the genetic sequences of SARS-CoV, SARS-CoV-2, and the two Korean bat CoV strains 16BO133 and B15-21, to estimate the likelihood of an interaction between the Korean bat CoVs and the human angiotensin-converting enzyme 2 (ACE2) receptor. Methods: The phylogenetic analysis was conducted with the maximum-likelihood (ML) method using MEGA 7 software. The Korean bat CoVs receptor binding domain (RBD) of the spike protein was analyzed by comparative homology modeling using the SWISS-MODEL server. The binding energies of the complexes were calculated using PRODIGY and MM/GBGA. Results: Phylogenetic analyses of the entire RNA-dependent RNA polymerase, spike regions, and the complete genome revealed that the Korean CoVs, along with SARS-CoV and SARS-CoV-2, belong to the subgenus Sarbecovirus, within BetaCoVs. However, the two Korean CoVs were distinct from SARS-CoV-2. Specifically, the spike gene of the Korean CoVs, which is involved in host infection, differed from that of SARS-CoV-2, showing only 66.8%-67.0% nucleotide homology and presented deletions within the RBD, particularly within regions critical for cross-species transmission and that mediate interaction with ACE2. Binding free energy calculation revealed that the binding affinity of Korean bat CoV RBD to hACE2 was drastically lower than that of SARS-CoV and SARS-CoV-2. Conclusions: These results suggest that Korean bat CoVs are unlikely to bind to the human ACE2 receptor.
Purpose: In Japan, the data on severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) antibody titers after the booster dose of the coronavirus disease 2019 (COVID-19) vaccine are insufficient. The aim of this study is to evaluate changes in SARS-CoV-2 antibody titers before, 1, 3, and 6 months after the booster dose of the BNT162b2 COVID-19 vaccine among health care workers. Materials and Methods: A total of 268 participants who received the booster dose of the BNT162b2 vaccine were analyzed. SARS-CoV-2 antibody titers were measured before (baseline) and at 1, 3, and 6 months after the booster dose. Factors associated with changes in SARS-CoV-2 antibody titers at 1, 3, and 6 months were analyzed. Cutoff values at baseline were calculated to prevent infection of the omicron variant of COVID-19. Results: The SARS-CoV-2 antibody titers at baseline, and 1, 3, and 6 months were 1,018.3 AU/mL, 21,396.5 AU/mL, 13,704.6 AU/mL, and 8,155.6 AU/mL, respectively. Factors associated with changes in SARS-CoV-2 antibody titers at 1 month were age and SARS-CoV-2 antibody titers at baseline, whereas changes in SARS-CoV-2 antibody titers at 3 and 6 months were associated with the SARS-CoV-2 antibody titers at 1 month. The cutoff values of the SARS-CoV-2 antibody titers at baseline were 515.4 AU/mL and 13,602.7 AU/mL at baseline and 1 month after the booster dose, respectively. Conclusion: This study showed that SARS-CoV-2 antibody titers increase rapidly at 1 month after the booster dose of the BNT162b2 vaccine and begin to decrease from 1 to 6 months. Hence, another booster may be needed as soon as possible to prevent infection.
Guo, Xi;Geng, Peng;Wang, Quan;Cao, Boyang;Liu, Bin
Journal of Microbiology and Biotechnology
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v.24
no.10
/
pp.1445-1454
/
2014
Severe acute respiratory syndrome (SARS), a disease that spread widely in the world during late 2002 to 2004, severely threatened public health. Although there have been no reported infections since 2004, the extremely pathogenic SARS coronavirus (SARS-CoV), as the causative agent of SARS, has recently been identified in animals, showing the potential for the re-emergence of this disease. Previous studies showed that 27 single nucleotide polymorphism (SNP) mutations among the spike (S) gene of this virus are correlated closely with the SARS pathogenicity and epidemicity. We have developed a SNP DNA microarray in order to detect and genotype these SNPs, and to obtain related information on the pathogenicity and epidemicity of a given strain. The microarray was hybridized with PCR products amplified from cDNAs obtained from different SARS-CoV strains. We were able to detect 24 SNPs and determine the type of a given strain. The hybridization profile showed that 19 samples were detected and genotyped correctly by using our microarray, with 100% accuracy. Our microarray provides a novel method for the detection and epidemiological surveillance of SARS-CoV.
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