title: [Prokaryotic expression and immunogenicity analysis of the chimeric HBcAg containing APP beta cleavage site peptide and Aβ(1-15);].
abstract: OBJECTIVE
  To construct the recombinant prokaryotic expression plasmid pET/c-ABCSP-Aβ(15-c);,  and evaluate the immunogenicity of the fusion protein expressed in  E.coli.
  
  
  METHODS
  The gene fragment HBc88-144 was amplified by PCR and subcloned to pUC19. The  APP beta cleavage site  peptide(ABCSP) and  Aβ(1-15); gene(ABCSP-Aβ(15);)  was amplified by PCR and inserted downstream of HBc1-71 in pGEMEX/c1-71. After restriction enzyme digestion,  c1-17-ABCSP-Aβ(15); were connected with  HBc88-144,  yielding the recombinant gene c-ABCSP-Aβ(15-c);. c-ABCSP-Aβ(15-c); gene was  subcloned into pET-28a(+).The fusion protein expressed in transformed E.coli BL21 was induced with IPTG and analyzed by SDS-PAGE. The virus-like particles (VLP) formed by fusion protein was observed with Transmission Electron Microscope (TEM). 4 Kunming (KM) mice received intraperitoneal injection (i.p) of fusion protein VLP. The antibody was detected by indirect ELISA.
  
  
  RESULTS
  The recombinant gene was confirmed by restriction enzyme digestion and DNA sequencing. After IPTG induction,  fusion protein was expressed and mainly existed in the sediment of the bacterial lysate. The expression level was 40% of all the proteins in the sediment. The fusion protein could form VLP. After 5 times of immunization,  the titer of anti-ABCSP and anti-Aβantibody in sera of KM mice reached up to 1:5 000 and 1:10 000 respectively,  while the anti-HBc antibody was undetectable.
  
  
  CONCLUSIONS
  Recombinant c-ABCSP-Aβ(15-c); gene can be expressed in E.coli. The expressed protein could form VLP and has a strong immunogenicity. This study lays the foundation for the study of AD genetic engineering vaccine.
authors.affiliation: Department of Human Anatomy and Histo/Embryology, Xi'an Jiaotong University, Xi'an, China. fenggaifeng@163.com
authors.forename: Gai-feng
authors.initials: GF
authors.lastname: Feng
authors.forename: Jun-yang
authors.initials: JY
authors.lastname: Wang
authors.forename: Hui
authors.initials: H
authors.lastname: Jin
authors.forename: Wei-xi
authors.initials: WX
authors.lastname: Wang
authors.forename: Yi-hua
authors.initials: YH
authors.lastname: Qian
authors.forename: Wei-na
authors.initials: WN
authors.lastname: Yang
authors.forename: Quan-ying
authors.initials: QY
authors.lastname: Wang
authors.forename: Guang-xiao
authors.initials: GX
authors.lastname: Yang
mesh_terms: D016229:Amyloid beta-Peptides / Q000235:genetics*; D000818:Animals; D002678:Chimera / Q000235:genetics; D005260:Female; D005822:Genetic Vectors / Q000235:genetics; D006512:Hepatitis B Core Antigens / Q000235:genetics*; D051379:Mice; D010446:Peptide Fragments / Q000235:genetics*; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000276:immunology* / Q000378:metabolism*; D058425:Vaccines, Virus-Like Particle / Q000235:genetics / Q000276:immunology* / Q000378:metabolism
chemical_list: D016229:Amyloid beta-Peptides; D006512:Hepatitis B Core Antigens; D010446:Peptide Fragments; D011993:Recombinant Fusion Proteins; D058425:Vaccines, Virus-Like Particle; C480626:amyloid beta-protein (1-15)