tetano
Editor, Senior Moderator
J Virol Methods. 2013 Sep 13. pii: S0166-0934(13)00387-X. doi: 10.1016/j.jviromet.2013.08.040. [Epub ahead of print]
Expression and purification of bioactive hemagglutinin protein of highly pathogenic avian influenza A (H5N1) in silkworm larvae.
Dong J, Harada M, Yoshida S, Kato Y, Murakawa A, Ogata M, Kato T, Usui T, Park EY.
Source
Department of Bioscience, Graduate School of Science and Technology.
Abstract
The hemagglutinin (HA) of avian influenza viruses plays a very important role in the infection of host cells. In this study, the HA gene of the highly pathogenic avian influenza H5N1 virus was cloned and expressed in silkworm larvae. The expressed recombinant HA (rHA) was purified using fetuin-agarose chromatography and Superdex 200 10/300 GL gel filtration chromatography, and the identity of purified rHA was confirmed by SDS-PAGE and Western blot. Approximately 500μg of purified rHA was obtained from a total of 30 silkworm larvae, suggesting the high efficiency of the silkworm expression system. The purified rHA bound to a rabbit polyclonal antibody against influenza A virus H5N1 (avian flu) HA, suggesting its antigenicity and potential application in vaccine development. Gel filtration chromatography showed that purified HA was present in the void volume fractions, indicating that rHA may form an oligomer. The rHA bound to poly{Neu5Acα2,3LacNAcβ-O[(CH2)5NHCO]2(CH2)5NH-/γ-PGA}, which mimics an avian type receptor, but did not bind to γ-polyglutamic acid or human type receptor mimic, poly{Neu5Acα2,6LacNAcβ-O[(CH2)5NHCO]2(CH2)5NH-/γ-PGA}, suggesting that it could be utilized as a blocking agent against infection by highly pathogenic influenza viruses.
Copyright ? 2013. Published by Elsevier B.V.
KEYWORDS:
Bacmid, Bombyx mori nucleopolyhedrovirus, Hemagglutinin, Influenza virus, Silkworm
PMID:
24041500
[PubMed - as supplied by publisher]
http://www.ncbi.nlm.nih.gov/pubmed/24041500
Expression and purification of bioactive hemagglutinin protein of highly pathogenic avian influenza A (H5N1) in silkworm larvae.
Dong J, Harada M, Yoshida S, Kato Y, Murakawa A, Ogata M, Kato T, Usui T, Park EY.
Source
Department of Bioscience, Graduate School of Science and Technology.
Abstract
The hemagglutinin (HA) of avian influenza viruses plays a very important role in the infection of host cells. In this study, the HA gene of the highly pathogenic avian influenza H5N1 virus was cloned and expressed in silkworm larvae. The expressed recombinant HA (rHA) was purified using fetuin-agarose chromatography and Superdex 200 10/300 GL gel filtration chromatography, and the identity of purified rHA was confirmed by SDS-PAGE and Western blot. Approximately 500μg of purified rHA was obtained from a total of 30 silkworm larvae, suggesting the high efficiency of the silkworm expression system. The purified rHA bound to a rabbit polyclonal antibody against influenza A virus H5N1 (avian flu) HA, suggesting its antigenicity and potential application in vaccine development. Gel filtration chromatography showed that purified HA was present in the void volume fractions, indicating that rHA may form an oligomer. The rHA bound to poly{Neu5Acα2,3LacNAcβ-O[(CH2)5NHCO]2(CH2)5NH-/γ-PGA}, which mimics an avian type receptor, but did not bind to γ-polyglutamic acid or human type receptor mimic, poly{Neu5Acα2,6LacNAcβ-O[(CH2)5NHCO]2(CH2)5NH-/γ-PGA}, suggesting that it could be utilized as a blocking agent against infection by highly pathogenic influenza viruses.
Copyright ? 2013. Published by Elsevier B.V.
KEYWORDS:
Bacmid, Bombyx mori nucleopolyhedrovirus, Hemagglutinin, Influenza virus, Silkworm
PMID:
24041500
[PubMed - as supplied by publisher]
http://www.ncbi.nlm.nih.gov/pubmed/24041500