title: Labeling of multiple HIV-1 proteins with the biarsenical-tetracysteine system.
abstract: Due to its small size and versatility, the biarsenical-tetracysteine system is an attractive way to label viral proteins for live cell imaging. This study describes the genetic labeling of the human immunodeficiency virus type 1 (HIV-1) structural proteins (matrix, capsid and nucleocapsid), enzymes (protease, reverse transcriptase, RNAse H and integrase) and envelope glycoprotein 120 with a tetracysteine tag in the context of a full-length virus. We measure the impact of these modifications on the natural virus infection and, most importantly, present the first infectious HIV-1 construct containing a fluorescently-labeled nucleocapsid protein. Furthermore, due to the high background levels normally associated with the labeling of tetracysteine-tagged proteins we have also optimized a metabolic labeling system that produces infectious virus containing the natural envelope glycoproteins and specifically labeled tetracysteine-tagged proteins that can easily be detected after virus infection of T-lymphocytes. This approach can be adapted to other viral systems for the visualization of the interplay between virus and host cell during infection.
authors.affiliation: Centre for Virology, Burnet Institute, Melbourne, Victoria, Australia.
authors.forename: Cândida F
authors.initials: CF
authors.lastname: Pereira
authors.forename: Paula C
authors.initials: PC
authors.lastname: Ellenberg
authors.forename: Kate L
authors.initials: KL
authors.lastname: Jones
authors.forename: Tara L
authors.initials: TL
authors.lastname: Fernandez
authors.forename: Redmond P
authors.initials: RP
authors.lastname: Smyth
authors.forename: David J
authors.initials: DJ
authors.lastname: Hawkes
authors.forename: Marcel
authors.initials: M
authors.lastname: Hijnen
authors.forename: Valérie
authors.initials: V
authors.lastname: Vivet-Boudou
authors.forename: Roland
authors.initials: R
authors.lastname: Marquet
authors.forename: Iain
authors.initials: I
authors.lastname: Johnson
authors.forename: Johnson
authors.initials: J
authors.lastname: Mak
mesh_terms: D020816:Amino Acid Motifs; D000595:Amino Acid Sequence; D001152:Arsenicals / Q000378:metabolism*; D003545:Cysteine; D057809:HEK293 Cells; D015497:HIV-1 / Q000201:enzymology / Q000235:genetics / Q000378:metabolism* / Q000472:pathogenicity; D006801:Humans; D008958:Models, Molecular; D008969:Molecular Sequence Data; D009154:Mutation; D011487:Protein Conformation; D013194:Staining and Labeling / Q000379:methods*; D013379:Substrate Specificity; D014764:Viral Proteins / Q000737:chemistry* / Q000378:metabolism*
chemical_list: D001152:Arsenicals; D014764:Viral Proteins; D003545:Cysteine