Opening and refolding of simian virus 40 and in vitro packaging of foreign DNA.
Simian virus 40 (SV40) can be disassembled under mild conditions by reducing disulfide bonds in the capsid and removing calcium ions. The nucleoprotein complexes formed, analyzed by electron microscopy, were circular and made up of 59 +/- 4 subunits, each with a diameter of about 10 nm. The complexes contained the viral DNA, histones, and the viral capsid proteins. The complexes had much-reduced infectivities compared with intact SV40. Addition of calcium ions to the disrupted virus caused the nucleoprotein complexes to refold into virus-like structures which sedimented at the same rate as intact SV40 and regained infectivity. Treatment of the disrupted SV40 with a high concentration of salt dissociated the viral proteins from the DNA. Lowering stepwise the salt concentration, removing the reducing agent, and adding calcium ions allowed structures to be reformed, and these structures sedimented, like SV40, at 240S and were infectious. The plaque-forming ability of the reconstituted particles was between that of the dissociated components and that of intact SV40. The addition of purified DNA of polyomavirus to the dissociated SV40 before the lowering of the salt concentration showed that virus-like structures could be formed from SV40 proteins and a foreign DNA.
Department of Virology, Swiss Institute for Experimental Cancer Research, Epalinges.
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MC
Colomar
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Degoumois-Sahli
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Beard
D002213:Capsid; D004261:DNA Replication; D004279:DNA, Viral; D005822:Genetic Vectors / Q000235:genetics*; D046911:Macromolecular Substances; D009690:Nucleic Acid Conformation; D011120:Polyomavirus / Q000235:genetics*; D011487:Protein Conformation; D013539:Simian virus 40 / Q000235:genetics* / Q000254:growth & development / Q000472:pathogenicity / Q000648:ultrastructure; D010948:Viral Plaque Assay; D014771:Virion / Q000235:genetics* / Q000254:growth & development / Q000472:pathogenicity / Q000648:ultrastructure; D014774:Virulence; D014779:Virus Replication
D004279:DNA, Viral; D046911:Macromolecular Substances