Large-scale purification and crystallization of adenovirus hexon.
This chapter provides a protocol for the large-scale purification of adenovirus type 2 and 5 virions and the soluble major coat protein hexon. The purified virus particles remain intact and are suitable for vector, vaccine, or structural studies and can also be used as seed stock for further rounds of infection. The hexon may be used to produce crystals suitable for high-resolution X-ray crystallographic studies. Briefly, virus is propagated in HeLa cell suspension cultures. The infected cells are lysed, virions and hexon are separated by centrifugation, and the protein is then further purified by anion exchange chromatography. The entire purification procedure takes approx 1 wk and typically yields 10(13) virus particles and 10-20 mg of highly purified hexon.
Rux
John J
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Burnett
Roger M
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D000256:Adenoviridae / Q000302:isolation & purification / Q000502:physiology; D036022:Capsid Proteins / Q000737:chemistry / Q000302:isolation & purification*; D002853:Chromatography, Liquid; D003460:Crystallization; D018360:Crystallography, X-Ray; D004591:Electrophoresis, Polyacrylamide Gel; D006367:HeLa Cells; D006801:Humans
D036022:Capsid Proteins; C018315:hexon capsid protein, Adenovirus