Abstract
The capsid protein is one of the three structural proteins of flaviviruses and is the building block of the nucleocapsid. It has also a predominant role in the replication of dengue virus. To obtain nucleocapsid-like particles from recombinant dengue-2 capsid protein produced in E. coli, a purification process using cation exchange chromatography was established. The purified protein exhibited a molecular mass corresponding to a dimer; therefore, similar to that reported for alphaviruses, an in vitro assembly reaction using single-stranded DNA was performed. In all cases, particles were obtained independently of the specificity and the length of the oligonucleotides used. The present work is the first report of in vitro assembly of the recombinant dengue capsid protein, which could constitute a powerful tool in the development of vaccine candidates.
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References
Gubler DJ (2002) Epidemic dengue/dengue hemorrhagic fever as a public health, social and economic problem in the 21st century. Trends Microbiol 10:100–103
Lindenbach BD, Rice CM (2003) Molecular biology of flaviviruses. Adv Virus Res 59:23–61
Ma L, Jones CT, Groesch TD, Kuhn RJ, Post CB (2004) Solution structure of dengue virus capsid protein reveals another fold. Proc Natl Acad Sci USA 101:3414–3419
Lobigs M (1993) Flavivirus premembrane protein cleavage and spike heterodimer secretion require the function of the viral proteinase NS3. Proc Natl Acad Sci U SA 90:6218–6222
Dokland T, Walsh M, Mackenzie JM, Khromykh AA, Kim-Huey E, Wang S (2004) West Nile virus core protein: tetramer structure and ribbon formation. Structure 12:1157–1163
Kuhn RJ, Zhang W, Rossmann MG, Pletnev SV, Corver J, Lenches E, Jones CT, Mukhopadhyay S, Chipman PR, Strauss EG, Baker TS, Strauss JH (2002) Structure of dengue virus: implications for flavivirus organization, maturation, and fusion. Cell 108:717–725
Mukhopadhyay S, Kim BS, Chipman PR, Rossmann MG, Kuhn RJ (2003) Structure of West Nile virus. Science 302:248
Ferlenghi I, Clarke M, Ruttan T, Allison SL, Schalich J, Heinz FX, Harrison SC, Rey FA, Fuller SD (2001) Molecular organization of a recombinant subviral particle from tick-borne encephalitis virus. Mol Cell 7:593–602
Kummerer BM, Rice CM (2002) Mutations in the yellow fever virus nonstructural protein NS2A selectively blocks production of infectious particles. J Virol 76:4773–4784
Jones CT, Ma L, Burgner JW, Groesch TD, Post CB, Kuhn RJ (2003) Flavivirus capsid is a dimeric alpha-helical protein. J Virol 77:7143–7149
Kiermayr S, Kofler MR, Mandl WC, Messner P, Heinz F (2004) Isolation of capsid protein dimers from the tick-borne encephalitis flavivirus and in vitro assembly of capsid-like particles. J Virol 78:8078–8084
Lazo L, Hermida L, Zulueta A, Sánchez J, López C, Silva R, Guillén G, Guzmán MG (2007) A recombinant capsid protein from dengue-2 induces protection in mice against homologous virus. Vaccine 25:1064–1070
Falcón V (1999) Ultrastructural and immunocytochemical evidences of core-particle formation in the methylotrophic Pichia pastoris yeast when expressing HCV structural proteins. Tissue Cell 31:117–125
Wengler G, Wengler G, Boege U, Wahn K (1984) Establishment and analysis of a system which allows assembly and disassembly of alphavirus core-like particles under physiological conditions in vitro. Virology 132:401–412
Acosta-Rivero N, Alvarez-Obregón JC, Musacchio A, Falcón V, Dueñas-Carrera S, Marante J, Menéndez I, Morales J (2002) In vitro self-assembled HCV core virus-like particles induce a strong antibody immune response in sheep. Biochem Biophys Res Commun 290:300–304
Acosta-Rivero N, Rodríguez A, Musacchio A, Falcón V, Suarez V, Martinez G, Guerra I, Paz-Lago D, Morera Y, de la Rosa MC, Morales-Grillo J, Dueñas-Carrera S (2004) In vitro assembly into virus-like particles is an intrinsic quality of Pichia pastoris derived HCV core protein. Biochem Biophys Res Commun 325:68–74
Tleugabulova D, Falcón V, Pentón E (1998) Evidence for the denaturation of recombinant hepatitis surface antigen on aluminum hydroxide gel. J Chromatogr Biomed Sci Appl 720:153–163
Goicochea NL, De M, Rotello VM, Mukhopadhyay S, Dragnea B (2007) Core-like particles of an enveloped animal virus can self-assemble efficiently on artificial templates. Nano Lett 7:2281–2290
Lobaina Y, Palenzuela D, Pichardo D, Muzio V, Guillen G, Aguilar JC (2005) Immunological characterization of two hepatitis B core antigen variants and their immunoenhancing effect on co-delivered hepatitis B surface antigen. Mol Immunol 42:289–294
Acknowledgment
The authors thank to Dr. Ricardo Silva (CIGB, Cuba) and Dr. Eduardo Martínez (CNC, Cuba) for the revision of the manuscript.
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López, C., Gil, L., Lazo, L. et al. In vitro assembly of nucleocapsid-like particles from purified recombinant capsid protein of dengue-2 virus. Arch Virol 154, 695–698 (2009). https://doi.org/10.1007/s00705-009-0350-8
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DOI: https://doi.org/10.1007/s00705-009-0350-8