Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:2.7.1.21 (thymidine kinase)
 7,561 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Protein p54, one of the most antigenic structural African swine fever virus (ASFV) proteins, has been localized by immuno-electron microscopy in the replication factories of infected cells, mainly associated with membranes and immature virus particles. Attempts to inactivate the p54 gene from ASFV by targeted insertion of beta-galactosidase selection marker was uniformly unsuccessful, suggesting that this gene is essential for virus viability. To demonstrate that, we inserted in the TK (thymidine kinase) locus of the virus a construction containing a second copy of the p54 gene and beta-glucuronidase selection marker under the control of p54 and p73 promoters, respectively. Virus mutant clones expressing a second copy of p54 and beta-glucuronidase were used to achieve deletion mutants of the original copy of the gene. Virus mutants expressing only the second inserted copy of p54 and the two selection markers mentioned above were successfully obtained. Therefore, we have demonstrated that the p54 gene product plays an essential role in virus growth, characterizing for the first time in ASFV an essential virus gene.
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PMID:The structural protein p54 is essential for African swine fever virus viability. 872 12

In the present work we have studied the feasibility of introducing foreign epitopes into the African swine fever virus (ASFV) particle by genetic manipulation of the virus. For this purpose, we developed specific transfer vectors containing the gene encoding for the highly antigenic structural ASFV protein p54 in which foreign sequences were introduced. DNA sequences encoding continuous linear epitopes, the antigenic site A from foot-and-mouth disease virus (FMDV) VP1 protein and the DA3 antigenic determinant from transmissible gastroenteritis coronavirus (TGEV) nucleoprotein N, were separately cloned into the p54 gene, in a region encoding a non-essential domain of the protein. Chimeric p54 genes were inserted by homologous recombination into the thymidine kinase (TK) locus of ASFV genome. The resulting recombinant viruses efficiently expressed both chimeric proteins under transcriptional control of the p54 promoter, and the chimeric gene products were recognized by antibodies to both p54 and foreign epitopes. The modified p54 proteins were also found in the viral particles and complemented the function of the wild-type p54, since deletion of the p54 gene from recombinant viruses did not affected virus replication in Vero cells. This work demonstrates for the first time the feasibility of incorporating foreign amino acid sequences (up to 18 residues) into a protein component of the ASFV particle without affecting virus viability.
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PMID:Design and construction of African swine fever virus chimeras incorporating foreign viral epitopes. 1048 37

Protective immunity to African swine fever virus (ASFV) may involve a combination of both serological and cellular mechanisms. This work is focused on the identification of the possible relevant serological immunodeterminants of immunity. Thus, 14 serological immunodeterminants of ASFV have been characterized by exhaustive screening of a representative lambda phage cDNA expression library of the tissue culture-adapted Ba71V strain of ASFV. The library was constructed using RNA extracted from Vero cells infected for 3, 6, 9 and 12 h. A total of 150 clones was selected arbitrarily by antibody screening of the library with a polyclonal antiserum from a domestic pig surviving infection with the virulent Malta isolate of ASFV. Sequencing of these clones permitted identification of 14 independent viral proteins that stimulated an antibody response. These included six proteins encoded by previously unassigned open reading frames (ORFs) (B602L, C44L, CP312R, E184L, K145R and K205R) as well as some of the more well-studied structural (A104R, p10, p32, p54 and p73) and non-structural proteins (RNA reductase, DNA ligase and thymidine kinase). Immunogenicity of these proteins was confirmed by demonstrating the corresponding antibodies in sera from pigs infected either with the Malta isolate or with the OURT88/3-OURT88/1 isolate combination. Furthermore, the majority of these ORFs were also recognized by immune antiserum from the natural host, the bush pig, following secondary challenge with the virulent Malawi (SINT90/1) isolate of ASFV. Thus, it is possible that some of these determinants may be important in protection against virus infection.
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PMID:Identification of the principal serological immunodeterminants of African swine fever virus by screening a virus cDNA library with antibody. 1202 48

The protective immune response to African swine fever virus (ASFV) includes both cellular and serological components. In this study, the role of antibodies in the pathogenicity and diagnosis of African swine fever (ASF) was explored. Accordingly, total and Ig isotype antibody responses against the 12 viral proteins previously demonstrated to be the main targets of serological immunity were evaluated in longitudinally collected sera from pigs infected experimentally with the non-pathogenic ASFV/NH/P68 isolate. Strong total IgG antibody responses were observed against viral proteins E183L/p54, K205R/'unassigned', A104R/histone-like and B602L/'unassigned'; therefore, IgM, IgG1 and IgG2 responses to these proteins were also determined. One protein stimulating IgM (K205R) may have practical potential for the detection of recently infected animals. There was a clear trend towards an IgG1 response to all of the proteins. This may reflect a dominant Th2-controlled immune response. In order to identify possible correlations between these serological responses and the pathogenesis of ASF, total IgG responses to the 12 recombinant proteins were compared in asymptomatic and chronically infected animals. For the proteins NP419L/DNA ligase, CP312R, B646L/p73, K196R/thymidine kinase and K205R, the antibody titres were significantly higher in animals developing lesions. One exception was the antibody response to the A104R/histone-like protein, which was higher in asymptomatic than in chronically infected pigs, suggesting that antibodies against this protein might be an indicator of an effective immune response or that this response is somehow involved in protection.
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PMID:Systematic analysis of longitudinal serological responses of pigs infected experimentally with African swine fever virus. 1769 51

The central variable region (CVR) within the B602L gene of the African swine fever virus (ASFV) is highly polymorphic within the 23 ASFV genotypes defined by sequencing of the C-terminal end of the p72 locus. Sequencing the p54 gene further discriminates ASFV genotypes that are conserved at the p72 locus. Variation in the thymidine kinase locus is a novel additional tool for ASFV genotyping whose application for this purpose is described for the first time herein. We evaluated genetic variation at these four polymorphic loci in 39 ASFV isolates obtained from outbreaks in Kenya and a region of Eastern Uganda between 2011 and 2013. Analysis of the p72 and p54 loci revealed high genetic conservation among these isolates; all clustered within p72 genotype IX and were similar to isolates associated with earlier outbreaks in East Africa. The thymidine kinase gene of the Kenyan isolates in this study were distinct relative to Southern African isolates and synonymous substitutions were observed among viruses from central Kenya. Analysis of the CVR within the B602L gene revealed two previously unknown polymorphisms that were restricted to Western Kenya and Eastern Uganda. A novel variant was revealed within CVR subgroup XXIV and a novel CVR subgroup XXIVa that contains tetrameric repeat F which has previously only been associated with p72 genotype I, was also identified for the first time in East Africa. Phylogeographic analysis of isolates based on CVR polymorphisms revealed rapid evolution and dissemination of variants present within ASFV genotype IX in East Africa.
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PMID:Multi-locus sequence typing of African swine fever viruses from endemic regions of Kenya and Eastern Uganda (2011-2013) reveals rapid B602L central variable region evolution. 2914 87