Quantitative analysis of the yield of avian H7 influenza virus haemagglutinin protein produced in silkworm pupae with the use of the codon-optimized DNA: A possible oral vaccine

Autor: Kenichi Maegawa, Sayaka Matsuda, Reiko Nerome, Shigeo Sugita, Kazumichi Kuroda, Kuniaki Nerome, Kazunori Kawasaki
Rok vydání: 2017
Předmět:
0301 basic medicine
Administration
Oral
Gene Expression
Hemagglutinin Glycoproteins
Influenza Virus
Antibodies
Viral
medicine.disease_cause
Virus
law.invention
Mice
03 medical and health sciences
chemistry.chemical_compound
0302 clinical medicine
Orthomyxoviridae Infections
Antigen
law
medicine
Animals
Amino Acid Sequence
Vaccines
Virus-Like Particle
030212 general & internal medicine
Codon
Vaccines
Synthetic
Influenza A Virus
H5N1 Subtype
General Veterinary
General Immunology and Microbiology
biology
fungi
Pupa
Public Health
Environmental and Occupational Health
Bombyx
Virology
Influenza A virus subtype H5N1
Blot
030104 developmental biology
Infectious Diseases
Immunization
chemistry
Influenza Vaccines
Larva
biology.protein
Recombinant DNA
Influenza A Virus
H7N1 Subtype
Molecular Medicine
Female
Antibody
Baculoviridae
Chickens
DNA
Zdroj: Vaccine. 35:738-746
ISSN: 0264-410X
DOI: 10.1016/j.vaccine.2016.12.058
Popis: In this study, we aimed to quantitatively compare the increased production of three H7 influenza virus-like particle (VLP) haemagglutinin (HA) with the use of a codon-optimized single HA gene in silkworm pupae. Recombinant baculovirus (Korea H7-BmNPV) could produce 0.40 million HA units per pupa, corresponding to 1832μg protein. The yield of the HA produced in larva was estimated to be approximately 0.31 million HA units per larva, and there were no significant differences between the three HA proteins. We could establish efficient recovery system of HA production in larvae and pupae with the use of three cycles sonication methods. Next, we compared yields of HA proteins from three different H7 and two H5 recombinant baculoviruses based on the amount of mRNA synthesized in BmN cells, suggesting that mRNA synthesis may be also a useful indicator for the production of HA. Based on HA titres from four recombinants, the yield of HA had a great influence on the codon-optimized effect and the characteristics of the viral HA gene. The recombinant containing codon optimized HA DNA of A/tufted duck/Fukushima/16/2011 (H5N1) did produce more than one million HA units, although another recombinant including of the wild H5N1 strain failed to show HA activity. Electron microscopy revealed the presence of large VLP and small HA particle in the heavy and light fractions. The purified VLPs reacted with the authentic anti-H7 antibodies and the antibodies prepared after immunization with the VLP H7 antigen. Also H5 and H7VLPs could produce HI antibody in chickens and mice with oral immunization. The antibodies elicited with oral immunization were confirmed in fluorescent antibody analysis and western blotting in Korea H5-BmNPV and H7HA-BmNPV recombinant infected BmN cells. Taken together, these findings provided important insights into future oral vaccine development.
Databáze: OpenAIRE
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