title: High yields of stable and highly pure nucleocapsid proteins of different hantaviruses can be generated in the yeast Saccharomyces cerevisiae.
abstract: Recently, the high-level expression of authentic and hexahistidine (His)-tagged Puumala (strain Vranica/Hällnäs) hantavirus nucleocapsid protein derivatives in the yeast Saccharomyces cerevisiae has been reported [Dargeviciute et al., Vaccine, 20 (2002) 3523-3531]. Here we describe the expression of His-tagged nucleocapsid proteins of other Puumala virus strains (Sotkamo, Kazan) as well as Dobrava (strains Slovenia and Slovakia) and Hantaan (strain Fojnica) hantaviruses using the same system. All nucleocapsid proteins were expressed in the yeast S. cerevisiae at high levels. The nucleocapsid proteins can be easily purified by nickel chelate chromatography; the yield for all nucleocapsid proteins ranged from 0.5 to 1.5 mg per g wet weight of yeast cells. In general, long-term storage of all nucleocapsid proteins without degradation can be obtained by storage in PBS at -20 degrees C or lyophilization. The nucleocapsid protein of Puumala virus (strain Vranica/Hällnäs) was demonstrated to contain only traces of less than 10 pg nucleic acid contamination per 100 microg of protein. The yeast-expressed nucleocapsid proteins of Hantaan, Puumala and Dobrava viruses described here represent useful tools for serological hantavirus diagnostics and for vaccine development.
authors.affiliation: Institute of Biotechnology, V. Graiciuno 8, LT-2028 Vilnius, Lithuania. ausra@ibt.lt
authors.forename: Ausra
authors.initials: A
authors.lastname: Razanskiene
authors.forename: Jonas
authors.initials: J
authors.lastname: Schmidt
authors.forename: Astrid
authors.initials: A
authors.lastname: Geldmacher
authors.forename: Andreas
authors.initials: A
authors.lastname: Ritzi
authors.forename: Matthias
authors.initials: M
authors.lastname: Niedrig
authors.forename: Ake
authors.initials: A
authors.lastname: Lundkvist
authors.forename: Detlev H
authors.initials: DH
authors.lastname: Krüger
authors.forename: Helga
authors.initials: H
authors.lastname: Meisel
authors.forename: Kestutis
authors.initials: K
authors.lastname: Sasnauskas
authors.forename: Rainer
authors.initials: R
authors.lastname: Ulrich
mesh_terms: D000818:Animals; D003001:Cloning, Molecular / Q000379:methods; D006476:Orthohantavirus / Q000235:genetics* / Q000378:metabolism*; D019590:Nucleocapsid Proteins / Q000096:biosynthesis* / Q000235:genetics* / Q000276:immunology / Q000302:isolation & purification; D015202:Protein Engineering / Q000379:methods*; D011817:Rabbits; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000276:immunology; D012441:Saccharomyces cerevisiae / Q000235:genetics* / Q000378:metabolism*
chemical_list: D019590:Nucleocapsid Proteins; D011994:Recombinant Proteins