title: Purification of functional baculovirus particles from silkworm larval hemolymph and their use as nanoparticles for the detection of human prorenin receptor (PRR) binding.
abstract: BACKGROUND
  Baculovirus, which has a width of 40 nm and a length of 250-300 nm, can display functional peptides, receptors and antigens on its surface by their fusion with a baculovirus envelop protein, GP64. In addition, some transmembrane proteins can be displayed without GP64 fusion, using the native transmembrane domains of the baculovirus. We used this functionality to display human prorenin receptor fused with GFPuv (GFPuv-hPRR) on the surface of silkworm Bombyx mori nucleopolyhedrovirus (BmNPV) and then tested whether these baculovirus particles could be used to detect protein-protein interactions.
  
  
  RESULTS
  BmNPV displaying GFPuv-hPRR (BmNPV-GFPuv-hPRR) was purified from hemolymph by using Sephacryl S-1000 column chromatography in the presence of 0.01% Triton X-100. Its recovery was 86% and the final baculovirus particles number was 4.98×10⁸ pfu. Based on the results of enzyme-linked immunosorbent assay (ELISA), 3.1% of the total proteins in BmNPV-GFPuv-hPRR were GFPuv-hPRR. This value was similar to that calculated from the result of western blot by a densitometry (2.7%). To determine whether BmNPV-GFPuv-hPRR particles were bound to human prorenin, ELISA results were compared with those from ELISAs using protease negative BmNPV displaying β1,3-N-acetylglucosaminyltransferase 2 fused with the gene encoding GFPuv (GGT2) (BmNPV-CP--GGT2) particles, which do not display hPRR on their surfaces.
  
  
  CONCLUSIONS
  The display of on the surface of the BmNPV particles will be useful for the detection of protein-protein interactions and the screening of inhibitors and drugs in their roles as nanobioparticles.
authors.affiliation: Laboratory of Biotechnology, Faculty of Agriculture, Shizuoka University, 836 Ohya, Suruga-ku, Shizuoka 422-8529, Japan.
authors.forename: Tatsuya
authors.initials: T
authors.lastname: Kato
authors.forename: Fumiaki
authors.initials: F
authors.lastname: Suzuki
authors.forename: Enoch Y
authors.initials: EY
authors.lastname: Park
mesh_terms: D000818:Animals; D012831:Bombyx / Q000821:virology*; D019286:Endopeptidase K / Q000378:metabolism; D004797:Enzyme-Linked Immunosorbent Assay; D006458:Hemolymph / Q000378:metabolism / Q000821:virology*; D006801:Humans; D007814:Larva / Q000821:virology; D053758:Nanoparticles / Q000737:chemistry*; D017924:Nucleopolyhedroviruses / Q000302:isolation & purification*; D017830:Octoxynol / Q000378:metabolism; D011485:Protein Binding; D011956:Receptors, Cell Surface / Q000032:analysis*; D012083:Renin / Q000378:metabolism; D014771:Virion / Q000302:isolation & purification*; D000093342:Prorenin Receptor
chemical_list: D011956:Receptors, Cell Surface; D017830:Octoxynol; D019286:Endopeptidase K; D012083:Renin; D000093342:Prorenin Receptor