Cell-free assembly of a polyoma-like particle from empty capsids and DNA

Barr, Stephen M.; Keck, Konrad; Aposhian, H. Vasken

doi:10.1016/0042-6822(79)90124-7

Cited by 34 publications

(15 citation statements)

References 8 publications

Supporting

Mentioning

Contrasting

Unclassified

Order By: Relevance

“…Complexes of VP1 structures with DNA were performed by incubation of the mixture of DNA with 5‐fold molar excess of pseudocapsids in buffer B for 1 h at 37°C, followed by exposure to osmotic shock, according to a previously described procedure [20].…”

Section: Methodsmentioning

confidence: 99%

Interactions of heterologous DNA with polyomavirus major structural protein, VP1

et al. 1999

View full text Add to dashboard Cite

Abstract`Empty' polyomavirus pseudocapsids, self-assembled from the major structural protein VP1, bind DNA nonspecifically and can deliver it into the nuclei of mammalian cells for expression [Forstova è et al. (1995) Hum. Gene Ther. 6, 2973 06]. Formation of suitable VP1-DNA complexes appears to be the limiting step in this route of gene delivery. Here, the character of VP1-DNA interactions has been studied in detail. Electron microscopy revealed that VP1 pseudocapsids can create in vitro at least two types of interactions with double-stranded DNA: (i) highly stable complexes, requiring free DNA ends, where the DNA is partially encapsidated; and, (ii) weaker interactions of pseudocapsids with internal parts of the DNA chain.z 1999 Federation of European Biochemical Societies.

show abstract

Section: Methodsmentioning

confidence: 99%

Interactions of heterologous DNA with polyomavirus major structural protein, VP1

et al. 1999

View full text Add to dashboard Cite

show abstract

“…DNA packaging and pseudofection with pseudocapsids DNA was packaged into pseudocapsids according to the procedure of Barr et al 21 For in vitro packaging, approximately 1 g of circular or linear DNA, containing the reporter gene, was mixed with 30 g of pseudocapsids and incubated at 37°C for 10 min. The mixture was exposed to osmotic shock by dilution with four volumes of H 2 O.…”

Section: Purification Of Polyoma Virus Empty Pseudocapsidsmentioning

confidence: 99%

Enhancement by polylysine of transient, but not stable, expression of genes carried into cells by polyoma VP1 pseudocapsids

et al. 1998

View full text Add to dashboard Cite

Gene transfer to provide long-term expression of a therapolylysine, unencapsidated DNA was shown to be propeutic product, without introducing unwelcome genetic tected against DNase activity, but electron microscopy information, is a goal being sought for therapy of both her-(EM) revealed the formation of large mixed aggregates. editary and acquired diseases. Polyoma virus pseudocap-The addition of pseudocapsids to these aggregates, and sids, generated from a VP1-expressing recombinant bacumeasurement of mobilities of the complexes in CsCl equililovirus, lack viral DNA and have been successfully used to brum centrifugation, indicated that they contained negliintroduce small exogenous genes stably into cells in vitro gible amounts of VP1. For subsequent pseudofection by a process designated 'pseudofection'; although pseudoexperiments, DNA was complexed first with pseudocapcapsids protect only about 3 kbp of exogenous DNA, low sids, then polylysine was added. The latter did not appear efficiency transfer of a larger fragment (6.2 kbp) has been to displace pseudocapsids from DNA, and was found to observed. Here, expression of a 7.2 kbp plasmid (pCMV␤) increase the efficiency of short-term expression both in encoding the ␤-galactosidase gene was assessed to moniin vitro and in vivo experiments. Gene expression, anator not only efficiency, but the ability of pseudocapsids to lysed histochemically or by the polymerase chain reaction, transfer larger-sized DNA on their own, or in the presence revealed transcriptional activity of the input gene, with of the polycation, poly-L-lysine, added to protect nonexpression first diminishing, then stabilising over time. The encapsidated DNA. When complexed to pseudocapsids presence of pseudocapsids, in complexes with DNA with only, the efficiency of expression of the transferred ␤-or without polylysine, allowed for stable and persistent galactosidase gene (in human or rodent cells), although gene expression. low, appeared to stabilise with time. In the presence of

show abstract

“…Loading Barr et al (1979) described for the first time the production of polyoma like particles (PLP) [54]. These particles were composed of purified, empty capsids and polyoma DNA.…”

Section: Assemblymentioning

confidence: 99%

“…These particles were stable in high salt concentrations. The integrated double-stranded DNA was protected against pancreatic DNase degradation [55] and the molecular weight amounted to approximately 1,1x10 6 Dalton [54]. Polyoma virus pseudocapsids or as well referred to as virus like particles (VLP) just composed of VP1 are able to incorporate exogenous DNA till 3kbp [56].…”

Section: Assemblymentioning

confidence: 99%

Recombinant Virus Like Particles as Drug Delivery System

Georgens¹,

Weyermann²,

Zimmer

2005

CPB

View full text Add to dashboard Cite

The drug delivery system described here is based on a virus like particle consisting of the recombinant expressed major capsid protein of Polyomavirus, VP1. Polyoma, a murine virus belonging to the Papovaviridae, forms a non-enveloped icosahedral capsid. These capsids are organized as a double shell composed of three different proteins: VP1,VP2 and VP3. The outer shell of the vision is composed of 360 VP1 molecules arranged as 72 pentamers. These capsids have a diameter of about 50 nm. The VP1 protein acts as a major ligand for certain membrane receptors during virus infection. Furthermore, the N-terminus of the VP1 protein contains a DNA-binding domain and a nuclear localization sequence. The recombinant production of the VP1 protein offers a save way to obtain a highly purified, non pathogenic pharmaceutical excipient. Combining these aspects, VP1 proteins provide a targeting as well as a drug binding site when used as a save drug carrier for gene therapy. Current applications are also including oligonucleotides as well as small molecules as well as vaccines.

show abstract

Cell-free assembly of a polyoma-like particle from empty capsids and DNA

Cited by 34 publications

References 8 publications

Interactions of heterologous DNA with polyomavirus major structural protein, VP1

Interactions of heterologous DNA with polyomavirus major structural protein, VP1

Enhancement by polylysine of transient, but not stable, expression of genes carried into cells by polyoma VP1 pseudocapsids

Recombinant Virus Like Particles as Drug Delivery System

Contact Info

Product

Resources

About