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Research Support, Non-U.S. Gov't
- Phosphorylation of the nucleocapsid protein of Hantaan virus by casein kinase II
- Jeong-Joong Yoon , Yun-Tai Lee , Hin Chu , Seung-yeol Son , Manbok Kim
- J. Microbiol. 2015;53(5):343-347. Published online May 3, 2015
- DOI: https://doi.org/10.1007/s12275-015-5095-3
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- 2 Crossref
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Abstract
- Hantaanvirus (HTNV) is the prototype of the genus Hantavirus, which belongs to the family Bunyaviridae. Hantaviruses are carried and transmitted by rodents and are known to cause two serious disease syndromes in humans i.e., hemorrhagic fever with renal syndrome (HFRS) and the hantavirus pulmonary syndrome (HPS). HTNV is an enveloped virus that contains a tripartite genome consisting of three negative-sense RNA segments (L, M, S), and the S and M segment of HTNV, respectively, encode the viral nucleocapsid protein (NP) and envelope glycoproteins. Possible phosphorylation motifs of casein kinase II (CKII) and protein kinase C (PKC) were identified in HTNV NP through bioinformatics searches. Sucrose gradient SDS-PAGE analysis indicated that dephosphorylated HTNV NP migrated faster than non-dephosphorylated NP, suggesting that HTNV NP is phosphorylated in infected Vero E6 cells. Immunoblot anaylsis of HTNV particles with anti-phosphoserine antibody and anti-phosphothreonine antibody after immunoprecipitation showed that viral particles are readily phosphorylated at threonine residues. In vitro kinase assay further showed that HTNV NP is phosphorylated by CK II, but not by PKC. Full length or truncated HTNV NPs expressed in E. coli were phosphorylated in vitro by CKII suggesting that phosphorylation may occur in vivo at multiple sites. Site specific mutagenesis studies suggest that HTNV NP phosphorylation might occur at unknown sites excluding the site-directly mutagenized locations. Taken together, HTNV NP can be phosphorylated mainly at threonine residues in vivo by CK II treatment.
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Citations
Citations to this article as recorded by
- Protein kinase CK2: a potential therapeutic target for diverse human diseases
Christian Borgo, Claudio D’Amore, Stefania Sarno, Mauro Salvi, Maria Ruzzene
Signal Transduction and Targeted Therapy.2021;[Epub] CrossRef - Unique Interferon Pathway Regulation by the Andes Virus Nucleocapsid Protein Is Conferred by Phosphorylation of Serine 386
Matthew J. Simons, Elena E. Gorbunova, Erich R. Mackow, Susana López
Journal of Virology.2019;[Epub] CrossRef
- Protein kinase CK2: a potential therapeutic target for diverse human diseases
Validation Study
- Development and Validation of a Recombinant Nucleocapsid Protein-Based ELISA for Detection of the Antibody to Porcine Reproductive and Respiratory Syndrome Virus
- Jia-Qi Chu , Xu-Min Hu , Myung-Cheol Kim , Chang-Sik Park , Moo-Hyung Jun
- J. Microbiol. 2009;47(5):582-588. Published online October 24, 2009
- DOI: https://doi.org/10.1007/s12275-009-0033-x
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- 13 Scopus
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Abstract
- Three indirect enzyme-linked immunosorbent assays (iELISA) based on the North American like (NA-like), European like (EU-like) and co-expressed NA- and EU-like recombinant nucleocapsid proteins (N-protein) of porcine reproductive and respiratory syndrome virus (PRRSV) were validated for the detection of the antibodies in porcine sera. A total of 422 serum samples from unvaccinated pigs were tested. The cut-off value was optimized by a two-graph receiver operating characteristics analysis at a 95% confidence level. This assay was validated with Western blot analysis and IDEXX HerdChek™ ELISA. Cross-reactivity results showed that iELISA was PRRSV-specific. Repeatability tests revealed that the coefficients of variation of positive sera within and between runs were less than 10%. The results indicate that iELISA is simpler to produce and perform, time-saving and suitable for large scale surveys of PRRSV infection at low cost, and is potentially useful to evaluate the efficiency of various vaccines against PRRSV.
Journal Articles
- Nucleocapsid Amino Acids 211 to 254, in Particular, Tetrad Glutamines, are Essential for the Interaction Between the Nucleocapsid and Membrane Proteins of SARS-Associated Coronavirus
- Xiaonan Fang , Lin-Bai Ye , Yijuan Zhang , Baozong Li , Shanshan Li , Lingbao Kong , Yuhua Wang , Hong Zheng , Wei Wang , Zhenghui Wu
- J. Microbiol. 2006;44(5):577-580.
- DOI: https://doi.org/2437 [pii]
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Abstract
- GST pull-down assays were used to characterize the SARS-CoV membrane (M) and nucleocapsid (N) interaction, and it was found that the amino acids 211-254 of N protein were essential for this interaction. When tetrad glutamines (Q) were replaced with glutamic acids (E) at positions of 240-243 of the N protein, the interaction was disrupted.
- Analysis of Immune Responses Against Nucleocapsid Protein of the Hantaan Virus Elicited by Virus Infection or DNA Vaccination
- Gyu-Jin Woo , Eun-Young Chun , Keun Hee Kim , Wankee Kim
- J. Microbiol. 2005;43(6):537-545.
- DOI: https://doi.org/2292 [pii]
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Abstract
- Even though neutralizing antibodies against the Hantaan virus (HTNV) has been proven to be critical against viral infections, the cellular immune responses to HTNV are also assumed to be important for viral clearance. In this report, we have examined the cellular and humoral immune responses against the HTNV nucleocapsid protein (NP) elicited by virus infection or DNA vaccination. To examine the cellular immune response against HTNV NP, we used H-2Kb restricted T-cell epitopes of NP. The NP-specific CD8+ T cell response was analyzed using a 51Cr-release assay, intracellular cytokine staining assay, enzyme-linked immunospot assay and tetramer binding assay in C57BL/6 mice infected with HTNV. Using these methods, we found that HTNV infection elicited a strong NP-specific CD8+ T cell response at eight days after infection. We also found that several different methods to check the NP-specific CD8+ T cell response showed a very high correlation among analysis. In the case of DNA vaccination by plasmid encoding nucleocapsid gene, the NP-specific antibody response was elicited 2 ~ 4 weeks after immunization and maximized at 6~8 weeks. NP-specific CD8+ T cell response reached its peak 3 weeks after immunization. In a challenge test with the recombinant vaccinia virus expressing NP (rVV-HTNV-N), the rVV-HTNV-N titers in DNA vaccinated mice were decreased about 100-fold compared to the negative control mice.
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