title: Structural Insights into HIV-1 Vif-APOBEC3F Interaction.
abstract: The HIV-1 Vif protein inactivates the cellular antiviral cytidine deaminase APOBEC3F (A3F) in virus-infected cells by specifically targeting it for proteasomal degradation. Several studies identified Vif sequence motifs involved in A3F interaction, whereas a Vif-binding A3F interface was proposed based on our analysis of highly similar APOBEC3C (A3C). However, the structural mechanism of specific Vif-A3F recognition is still poorly understood. Here we report structural features of interaction interfaces for both HIV-1 Vif and A3F molecules. Alanine-scanning analysis of Vif revealed that six residues located within the conserved Vif F1-, F2-, and F3-box motifs are essential for both A3C and A3F degradation, and an additional four residues are uniquely required for A3F degradation. Modeling of the Vif structure on an HIV-1 Vif crystal structure revealed that three discontinuous flexible loops of Vif F1-, F2-, and F3-box motifs sterically cluster to form a flexible A3F interaction interface, which represents hydrophobic and positively charged surfaces. We found that the basic Vif interface patch (R17, E171, and R173) involved in the interactions with A3C and A3F differs. Furthermore, our crystal structure determination and extensive mutational analysis of the A3F C-terminal domain demonstrated that the A3F interface includes a unique acidic stretch (L291, A292, R293, and E324) crucial for Vif interaction, suggesting additional electrostatic complementarity to the Vif interface compared with the A3C interface. Taken together, these findings provide structural insights into the A3F-Vif interaction mechanism, which will provide an important basis for development of novel anti-HIV-1 drugs using cellular cytidine deaminases.
  
  
  OBJECTIVE
  HIV-1 Vif targets cellular antiviral APOBEC3F (A3F) enzyme for degradation. However, the details on the structural mechanism for specific A3F recognition remain unclear. This study reports structural features of interaction interfaces for both HIV-1 Vif and A3F molecules. Three discontinuous sequence motifs of Vif, F1, F2, and F3 boxes, assemble to form an A3F interaction interface. In addition, we determined a crystal structure of the wild-type A3F C-terminal domain responsible for the Vif interaction. These results demonstrated that both electrostatic and hydrophobic interactions are the key force driving Vif-A3F binding and that the Vif-A3F interfaces are larger than the Vif-A3C interfaces. These findings will allow us to determine the configurations of the Vif-A3F complex and to construct a structural model of the complex, which will provide an important basis for inhibitor development.
authors.affiliation: Clinical Research Center, National Hospital Organization Nagoya Medical Center, Nagoya, Aichi, Japan Department of Biotechnology, Nagoya University Graduate School of Engineering, Nagoya, Aichi, Japan.
authors.forename: Masaaki
authors.initials: M
authors.lastname: Nakashima
authors.affiliation: Clinical Research Center, National Hospital Organization Nagoya Medical Center, Nagoya, Aichi, Japan.
authors.forename: Hirotaka
authors.initials: H
authors.lastname: Ode
authors.affiliation: Synchrotron Radiation Research Center, Nagoya University, Nagoya, Aichi, Japan.
authors.forename: Takashi
authors.initials: T
authors.lastname: Kawamura
authors.affiliation: Clinical Research Center, National Hospital Organization Nagoya Medical Center, Nagoya, Aichi, Japan Department of Biotechnology, Nagoya University Graduate School of Engineering, Nagoya, Aichi, Japan.
authors.forename: Shingo
authors.initials: S
authors.lastname: Kitamura
authors.affiliation: Clinical Research Center, National Hospital Organization Nagoya Medical Center, Nagoya, Aichi, Japan.
authors.forename: Yuriko
authors.initials: Y
authors.lastname: Naganawa
authors.affiliation: Clinical Research Center, National Hospital Organization Nagoya Medical Center, Nagoya, Aichi, Japan Department of Biotechnology, Nagoya University Graduate School of Engineering, Nagoya, Aichi, Japan.
authors.forename: Hiroaki
authors.initials: H
authors.lastname: Awazu
authors.affiliation: Clinical Research Center, National Hospital Organization Nagoya Medical Center, Nagoya, Aichi, Japan Department of Biotechnology, Nagoya University Graduate School of Engineering, Nagoya, Aichi, Japan.
authors.forename: Shinya
authors.initials: S
authors.lastname: Tsuzuki
authors.affiliation: Clinical Research Center, National Hospital Organization Nagoya Medical Center, Nagoya, Aichi, Japan.
authors.forename: Kazuhiro
authors.initials: K
authors.lastname: Matsuoka
authors.affiliation: Clinical Research Center, National Hospital Organization Nagoya Medical Center, Nagoya, Aichi, Japan.
authors.forename: Michiko
authors.initials: M
authors.lastname: Nemoto
authors.affiliation: Clinical Research Center, National Hospital Organization Nagoya Medical Center, Nagoya, Aichi, Japan.
authors.forename: Atsuko
authors.initials: A
authors.lastname: Hachiya
authors.affiliation: Clinical Research Center, National Hospital Organization Nagoya Medical Center, Nagoya, Aichi, Japan Division of Basic Medicine, Nagoya University Graduate School of Medicine, Nagoya, Aichi, Japan.
authors.forename: Wataru
authors.initials: W
authors.lastname: Sugiura
authors.affiliation: Clinical Research Center, National Hospital Organization Nagoya Medical Center, Nagoya, Aichi, Japan.
authors.forename: Yoshiyuki
authors.initials: Y
authors.lastname: Yokomaku
authors.affiliation: Department of Biotechnology, Nagoya University Graduate School of Engineering, Nagoya, Aichi, Japan Synchrotron Radiation Research Center, Nagoya University, Nagoya, Aichi, Japan.
authors.forename: Nobuhisa
authors.initials: N
authors.lastname: Watanabe
authors.affiliation: Clinical Research Center, National Hospital Organization Nagoya Medical Center, Nagoya, Aichi, Japan Division of Basic Medicine, Nagoya University Graduate School of Medicine, Nagoya, Aichi, Japan iwataniy@nnh.hosp.go.jp.
authors.forename: Yasumasa
authors.initials: Y
authors.lastname: Iwatani
mesh_terms: D018360:Crystallography, X-Ray; D003564:Cytidine Deaminase / Q000737:chemistry / Q000378:metabolism; D043525:Cytosine Deaminase / Q000737:chemistry* / Q000378:metabolism*; D004252:DNA Mutational Analysis; D008958:Models, Molecular; D016297:Mutagenesis, Site-Directed; D011487:Protein Conformation; D025941:Protein Interaction Mapping; D059748:Proteolysis; D054320:vif Gene Products, Human Immunodeficiency Virus / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*
chemical_list: D054320:vif Gene Products, Human Immunodeficiency Virus; C514847:vif protein, Human immunodeficiency virus 1; C492906:APOBEC3F protein, human; D043525:Cytosine Deaminase; C495795:APOBEC3C protein, human; D003564:Cytidine Deaminase