Formation of Poliovirus-like Particles by Recombinant Baculoviruses Expressing the Individual VP0, VP3, and VP1 Proteins by Comparison to Particles Derived from the Expressed Poliovirus Polyprotein

Bräutigam, Sandra; Snezhkov, E. V.; Bishop, David H. L.

doi:10.1006/viro.1993.1067

Cited by 71 publications

(39 citation statements)

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“…When applied to the purification of poliovirus proteins from gel eluates, this technique produced fractions of high purity and permitted the identification of distinct capsid proteins recognised by a panel of poliovirus-specific T cell clones. Following the development of the technique, baculovirus expressed poliovirus proteins VP0 (precursor of VP2 and VP4), VP1 and VP3 became available (Br~iutigam et al, 1992). The pattern of responses of the T cell clones against the recombinant proteins confirmed the specificity established with latex microsphere-adsorbed material separated from viral lysates.…”

Section: Discussionmentioning

confidence: 84%

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Preparative separation of foreign antigens for highly efficient presentation to T cells in vitro

Katrak

Mahon

Jones

et al. 1992

Journal of Immunological Methods

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A method is described for the separation and purification of proteins from complex mixtures of foreign antigens in a form suitable for stimulating T cells in vitro. The technique involves electrophoretic separation of proteins followed by elution, concentration and adsorption of the polypeptide subunits to latex microspheres. Alternatively, where a specific antibody is available, proteins may be affinity-purified from a heterogeneous mixture of antigens, using antibody-coated latex microspheres. Nanogram quantities of protein coupled to latex were shown to be highly efficient stimulators of antigen-specific T cells as tested by in vitro proliferation and cytokine release assays. The utility of this technique was demonstrated using poliovirus capsid proteins separated by SDS-polyacrylamide gel electrophoresis (PAGE) and coupled to latex microspheres for specificity analysis of T cell clones. Antigen reactivity of the T cell clones was confirmed using recombinant baculoviruses expressing individual poliovirus proteins. Furthermore, recombinant proteins coupled to latex microspheres were used for efficient stimulation and in vitro propagation of T cell clones specific for the simian immunodeficiency virus (SIV) envelope (env) protein. Although the technique is illustrated in this report using viral antigens, it has also proved to be an efficient method for the separation of bacterial antigens in studies of polyclonal T cell responses to Bordetella pertussis antigens.

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Section: Discussionmentioning

confidence: 84%

“…The cDNA sequences encoding the three capsid proteins were cloned and expressed individually under the control of the polyhedrin promoter, as described by Br~iutigam et al (1992). Expression of these proteins was performed in infectedSpodoptera frugiperda cells.…”

Section: Antigensmentioning

confidence: 99%

Preparative separation of foreign antigens for highly efficient presentation to T cells in vitro

Katrak

Mahon

Jones

et al. 1992

Journal of Immunological Methods

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“…This is due the fact that for efficient assembly of a VLP the interacting capsid proteins must be expressed in the vicinity to each other, in other words in the same cell. Assembly of VLPs by processing of polyproteins have been achieved both for poliovirus 33 and for Infectious Bursal disease virus 34 using the baculovirus expression system. More complex assembly of multilayered, multiprotein VLPs have also been efficiently produced for the members of the Reoviridae.…”

Section: Vlps Of Structurally Complex Viral Capsids With Multiple Promentioning

confidence: 99%

Virus‑like particles as a vaccine delivery system: Myths and facts

Roy¹,

Noad²

2008

Human Vaccines

150

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Vaccines against viral disease have traditionally relied on attenuated virus strains or inactivation of infectious virus. Subunit vaccines based on viral proteins expressed in heterologous systems have been effective for some pathogens, but have often suffered from poor immunogenicity due to incorrect protein folding or modification. In this review we focus on a specific class of viral subunit vaccine that mimics the overall structure of virus particles and thus preserves the native antigenic conformation of the immunogenic proteins. These virus-like particles (VLPs) have been produced for a wide range of taxonomically and structurally distinct viruses, and have unique advantages in terms of safety and immunogenicity over previous approaches. With new VLP vaccines for papillomavirus beginning to reach the marketplace we argue that this technology has now 'come-of-age' and must be considered a viable vaccine strategy.

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“…Characterization of these events at the molecular level has been greatly facilitated by the understanding that viral structural proteins may contain all of the necessary information to dictate the assembly process. In fact, it was observed that coexpression of viral structural proteins would result in the formation of virus-like particles (VLPs) 1 in many different viral systems (1)(2)(3)(4)(5)(6)(7)(8)(9). In this study, we exploit the coronavirus VLP system to study the interaction between two structural proteins during coronavirus assembly and the implication of this interaction in the assembly and release of coronavirus particles.…”

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confidence: 99%

The Missing Link in Coronavirus Assembly

Lim

Liu

2001

Journal of Biological Chemistry

139

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One missing link in the coronavirus assembly is the physical interaction between two crucial structural proteins, the membrane (M) and envelope (E) proteins. In this study, we demonstrate that the coronavirus infectious bronchitis virus E can physically interact, via a putative peripheral domain, with M. Deletion of this domain resulted in a drastic reduction in the incorporation of M into virus-like particles. Immunofluorescent staining of cells coexpressing M and E supports that E interacts with M and relocates M to the same subcellular compartments that E resides in. E was retained in the pre-Golgi membranes, prior to being translocated to the Golgi apparatus and the secretory vesicles; M was observed to exhibit similar localization and translocation profiles as E when coexpressed with E. Deletion studies identified the C-terminal 6-residue RDKLYS as the endoplasmic reticulum retention signal of E, and site-directed mutagenesis of the ؊4 lysine residue to glutamine resulted in the accumulation of E in the Golgi apparatus. The third domain of E that plays a crucial role in virus budding is a putative transmembrane domain present at the N-terminal region, because deletion of the domain resulted in a free distribution of the mutant protein and in dysfunctional viral assembly.Morphogenesis and assembly of mammalian enveloped RNA viruses are complex processes. During these processes, the viral core, consisting of viral RNA and core proteins, becomes wrapped in a membranous structure (viral envelope) derived from host cell membranes to form virion particles. The assembly process, which is referred to as budding, usually occurs at either the plasma or intracellular membranes. The formation of viral core, envelope, and the assembly of virus particles would involve interaction between viral proteins and host membrane components, among viral structural proteins and between viral proteins and viral RNA. Characterization of these events at the molecular level has been greatly facilitated by the understanding that viral structural proteins may contain all of the necessary information to dictate the assembly process. In fact, it was observed that coexpression of viral structural proteins would result in the formation of virus-like particles (VLPs) 1 in many different viral systems (1-9). In this study, we exploit the coronavirus VLP system to study the interaction between two structural proteins during coronavirus assembly and the implication of this interaction in the assembly and release of coronavirus particles. Coronavirus is the largest RNA virus known so far. It has a positive-sense, single-strand RNA genome of 27-30 kilobases in length. Despite of the huge genome size, coronavirus typically contains four structural proteins, i.e. a type I spike (S) glycoprotein required for infectivity, a phosphorylated nucleocapsid (N) protein that interacts with the viral genome to form a helical core, a major type III integral membrane (M) protein, and a minor type III envelope (E) protein (10 -15). A fifth protein, the hemagglut...

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Formation of Poliovirus-like Particles by Recombinant Baculoviruses Expressing the Individual VP0, VP3, and VP1 Proteins by Comparison to Particles Derived from the Expressed Poliovirus Polyprotein

Cited by 71 publications

References 0 publications

Preparative separation of foreign antigens for highly efficient presentation to T cells in vitro

Preparative separation of foreign antigens for highly efficient presentation to T cells in vitro

Virus‑like particles as a vaccine delivery system: Myths and facts

The Missing Link in Coronavirus Assembly

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