Oligonucleotide and plasmid DNA packaging into polyoma VP1 virus-like particles expressed in Escherichia coli.
The drug delivery system described here is based on the properties of the capsoid or capsid-like structure resulting from the assembly of polyoma virus capsid protein VP1 expressed in Escherichia coli. The capsid protein VP1 was expressed as a fusion protein with a completely removable N-terminal His6 affinity tag. The pentameric morphology of the recombinant VP1 protein was confirmed by electron microscopy after affinity chromatography and factor Xa cleavage under conditions of low ionic strength. The self-assembly of VP1 capsoids can be induced from purified VP1 pentamers by increasing the ionic strength with (NH4)2SO4. These VP1 capsoid particles were packed in vitro with anti-sense oligonucleotides and plasmid DNA. The loading with DNA was pH-dependent. We observed the highest efficiency at pH 5. DNase I treatment of particles with encapsidated material showed that 37-55% of the bound oligonucleotides and fragments of 1.5-1.8 kb double-stranded DNA were protected against degradation.
Braun
H
H
Institute for Pharmaceutical Technology, University of Frankfurt, Biozentrum, Marie-Curie-Strasse 9, 60439 Frankfurt, Germany.
Boller
K
K
Löwer
J
J
Bertling
W M
WM
Zimmer
A
A
D001483:Base Sequence; D002213:Capsid / Q000737:chemistry* / Q000235:genetics* / Q000378:metabolism; D036022:Capsid Proteins; D003850:Deoxyribonuclease I / Q000737:chemistry / Q000378:metabolism; D004926:Escherichia coli / Q000235:genetics / Q000821:virology*; D005822:Genetic Vectors / Q000235:genetics; D006863:Hydrogen-Ion Concentration; D008969:Molecular Sequence Data; D009841:Oligonucleotides / Q000737:chemistry* / Q000235:genetics; D009997:Osmotic Pressure; D010957:Plasmids / Q000737:chemistry* / Q000235:genetics; D011120:Polyomavirus / Q000737:chemistry / Q000235:genetics; D011487:Protein Conformation; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D012492:Salts; D017421:Sequence Analysis
D036022:Capsid Proteins; D009841:Oligonucleotides; D011994:Recombinant Proteins; D012492:Salts; C062481:VP1 protein, polyomavirus; D003850:Deoxyribonuclease I