title: An effective and simplified DO-stat control strategy for production of rabies glycoprotein in Pichia pastoris.
abstract: The glycoprotein (G-protein) of rabies virus is responsible for viral attachment to the host cell surface and induces virus neutralization antibodies. In the present study, the G-protein gene of rabies virus CVS strain was cloned, sequenced and expressed in the yeast, Pichia pastoris, as a secreted protein, using a simplified DO-stat control feeding strategy. This strategy involves the addition of methanol when the dissolved oxygen (DO) level rises above the setpoint avoiding methanol accumulation and oxygen limitation. The G-protein expression was evaluated by SDS-PAGE, ELISA, and western blot assays. Like native G-protein, the recombinant G-protein was found reactive when it was challenged against specific antibodies. The data indicate that the recombinant G-protein can be easily expressed and isolated, and may be useful as a safe source in the production of diagnostic kits and subunit vaccines to prevent rabies.
authors.affiliation: CINDEFI, CONICET, La Plata, Argentina; Department of Virology, School of Veterinary Sciences, National University of La Plata, Buenos Aires, Argentina. Electronic address: lpicotto@fcv.unlp.edu.ar.
authors.forename: L D
authors.initials: LD
authors.lastname: Picotto
authors.affiliation: Department of Virology, School of Veterinary Sciences, National University of La Plata, Buenos Aires, Argentina.
authors.forename: G H
authors.initials: GH
authors.lastname: Sguazza
authors.affiliation: Department of Virology, School of Veterinary Sciences, National University of La Plata, Buenos Aires, Argentina.
authors.forename: M A
authors.initials: MA
authors.lastname: Tizzano
authors.affiliation: Department of Virology, School of Veterinary Sciences, National University of La Plata, Buenos Aires, Argentina; Scientific Research Commission, Buenos Aires Province (CIC-PBA), Argentina.
authors.forename: C M
authors.initials: CM
authors.lastname: Galosi
authors.affiliation: CINDEFI, CONICET, La Plata, Argentina; Department of Biotechnology, School of Exact Sciences, National University of La Plata, Buenos Aires, Argentina.
authors.forename: S F
authors.initials: SF
authors.lastname: Cavalitto
authors.affiliation: Department of Virology, School of Veterinary Sciences, National University of La Plata, Buenos Aires, Argentina.
authors.forename: M R
authors.initials: MR
authors.lastname: Pecoraro
mesh_terms: D010843:Pichia / Q000235:genetics / Q000378:metabolism*; D011820:Rabies virus / Q000235:genetics*; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000737:chemistry; D014759:Viral Envelope Proteins / Q000096:biosynthesis / Q000737:chemistry / Q000235:genetics
chemical_list: D011994:Recombinant Proteins; D014759:Viral Envelope Proteins
keywords: DO-stat; Glycoprotein; Pichia pastoris; Rabies virus