title: Structural basis for the inhibition of HIV-1 Nef by a high-affinity binding single-domain antibody.
abstract: BACKGROUND
  The HIV-1 Nef protein is essential for AIDS pathogenesis by its interaction with host cell surface receptors and signaling factors. Despite its critical role as a virulence factor Nef is not targeted by current antiviral strategies.
  
  
  RESULTS
  We have determined the crystal structure of the complex formed by a camelid single-domain antibody fragment, termed sdAb19, bound to HIV-1 Nef together with a stabilizing SH3 domain. sdAb19 forms a stoichiometric 1:1 complex with Nef and binds to a conformationally conserved surface at the C-terminus of Nef that overlaps with functionally important interaction sites involved in Nef-induced perturbations of signaling and trafficking pathways. The antibody fragment binds Nef with low nanomolar affinity, which could be attenuated to micromolar affinity range by site-directed mutagenesis of key interaction residues in sdAb19. Fusion of the SH3 domain to sdAb19, termed Neffin, leads to a significantly increased affinity for Nef and formation of a stoichiometric 2:2 Nef-Neffin complex. The 19 kDa Neffin protein inhibits all functions of Nef as CD4 and MHC-I downregulation, association with Pak2, and the increase in virus infectivity and replication.
  
  
  CONCLUSIONS
  Together, sdAb19 and Neffin thus represent efficient tools for the rational development of antiviral strategies against HIV-1 Nef.
authors.forename: Sebastian
authors.initials: S
authors.lastname: Lülf
authors.forename: Julie
authors.initials: J
authors.lastname: Matz
authors.forename: Marie-Christine
authors.initials: MC
authors.lastname: Rouyez
authors.forename: Annika
authors.initials: A
authors.lastname: Järviluoma
authors.forename: Kalle
authors.initials: K
authors.lastname: Saksela
authors.forename: Serge
authors.initials: S
authors.lastname: Benichou
authors.affiliation: Center of Advanced European Studies and Research, Group Physical Biochemistry, Bonn, Germany. matthias.geyer@caesar.de.
authors.forename: Matthias
authors.initials: M
authors.lastname: Geyer
mesh_terms: D015483:HIV Antibodies / Q000737:chemistry* / Q000276:immunology*; D015497:HIV-1 / Q000737:chemistry / Q000276:immunology; D006801:Humans; D011485:Protein Binding; D011487:Protein Conformation; D061905:Single-Domain Antibodies / Q000737:chemistry* / Q000276:immunology*; D054311:nef Gene Products, Human Immunodeficiency Virus / Q000737:chemistry* / Q000276:immunology*
chemical_list: D015483:HIV Antibodies; D061905:Single-Domain Antibodies; D054311:nef Gene Products, Human Immunodeficiency Virus; C514837:nef protein, Human immunodeficiency virus 1