Polymerization of murine recombinant prion protein in nucleic acid solution.
Recombinant prion protein has been used earlier to understand the structural properties of cellular prion protein PrP(C) and to understand conformational change of PrP(C) to its isoform, PrP(Sc) which is believed to be responsible for the prion disease. Here we report that murine recombinant prion protein, MoPrP(C) polymerizes in the presence of nucleic acid. The aggregation process and the properties of the aggregates have been monitored by physical, biochemical and ultrastructural studies. An increase in the turbidity at 0,90 degrees light scattering is observed when the protein is added to nucleic acid. An increase in the fluorescence of anilino naphthalene sulfonic acid dye (ANS) accompanying a blue shift in its emission maxima is observed when the aggregate obtained from prion protein and DNA reaction is added to it. The kinetics of the increase of the ANS fluorescence during aggregation process show lag periods which depend linearly on the nucleic acid concentration but show a biphasic dependence on the protein concentration. The change in the fluorescence properties of the dye in the presence of the aggregates obtained in the present study and in the presence of the protein PrP 27-30 amyloid isolated in vivo reported in literature are similar. The dye Congo Red binds to the aggregates resulting from the aggregation reaction.The ultrastructural analysis revealed polymeric structures with amyloid like morphologies and smaller oligomeric structures. In addition, condensed nucleic acid structures are also observed which are morphologically different from histone induced condensed nucleic acid structures but are similar to Human Immunodeficiency Virus-1 nucleocapsid protein, NCp7, induced nucleic acid structures. The aggregates show resistance to degradation by proteinase K treatment. Charge neutralization resulting from the MoPrP(C)-DNA interaction and accompanying structural changes in the molecules may explain the observed effects.
Nandi
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INRA, Pathologie Infectieuse et Immunologie, Nouzilly, France.
Leclerc
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E
D000817:Anilino Naphthalenesulfonates; D000818:Animals; D004396:Coloring Agents; D003224:Congo Red; D004247:DNA / Q000737:chemistry*; D005456:Fluorescent Dyes; D007700:Kinetics; D051379:Mice; D008854:Microscopy, Electron; D011108:Polymers / Q000737:chemistry; D018621:PrPC Proteins / Q000737:chemistry*; D011994:Recombinant Proteins / Q000737:chemistry; D013050:Spectrometry, Fluorescence
D000817:Anilino Naphthalenesulfonates; D004396:Coloring Agents; D005456:Fluorescent Dyes; D011108:Polymers; D018621:PrPC Proteins; D011994:Recombinant Proteins; D003224:Congo Red; D004247:DNA