title: Crystal structure of an antiviral ankyrin targeting the HIV-1 capsid and molecular modeling of the ankyrin-capsid complex.
abstract: Ankyrins are cellular repeat proteins, which can be genetically modified to randomize amino-acid residues located at defined positions in each repeat unit, and thus create a potential binding surface adaptable to macromolecular ligands. From a phage-display library of artificial ankyrins, we have isolated Ank(GAG)1D4, a trimodular ankyrin which binds to the HIV-1 capsid protein N-terminal domain (NTD(CA)) and has an antiviral effect at the late steps of the virus life cycle. In this study, the determinants of the Ank(GAG)1D4-NTD(CA) interaction were analyzed using peptide scanning in competition ELISA, capsid mutagenesis, ankyrin crystallography and molecular modeling. We determined the Ank(GAG)1D4 structure at 2.2 Å resolution, and used the crystal structure in molecular docking with a homology model of HIV-1 capsid. Our results indicated that NTD(CA) alpha-helices H1 and H7 could mediate the formation of the capsid-Ank(GAG)1D4 binary complex, but the interaction involving H7 was predicted to be more stable than with H1. Arginine-18 (R18) in H1, and R132 and R143 in H7 were found to be the key players of the Ank(GAG)1D4-NTD(CA) interaction. This was confirmed by R-to-A mutagenesis of NTD(CA), and by sequence analysis of trimodular ankyrins negative for capsid binding. In Ank(GAG)1D4, major interactors common to H1 and H7 were found to be S45, Y56, R89, K122 and K123. Collectively, our ankyrin-capsid binding analysis implied a significant degree of flexibility within the NTD(CA) domain of the HIV-1 capsid protein, and provided some clues for the design of new antivirals targeting the capsid protein and viral assembly.
authors.affiliation: Division of Clinical Immunology, Department of Medical Technology, Faculty of Associated Medical Sciences, Chiang Mai University, Chiang Mai, 50200, Thailand.
authors.forename: Warachai
authors.initials: W
authors.lastname: Praditwongwan
authors.forename: Phimonphan
authors.initials: P
authors.lastname: Chuankhayan
authors.forename: Somphot
authors.initials: S
authors.lastname: Saoin
authors.forename: Tanchanok
authors.initials: T
authors.lastname: Wisitponchai
authors.forename: Vannajan Sanghiran
authors.initials: VS
authors.lastname: Lee
authors.forename: Sawitree
authors.initials: S
authors.lastname: Nangola
authors.forename: Saw See
authors.initials: SS
authors.lastname: Hong
authors.forename: Philippe
authors.initials: P
authors.lastname: Minard
authors.forename: Pierre
authors.initials: P
authors.lastname: Boulanger
authors.forename: Chun-Jung
authors.initials: CJ
authors.lastname: Chen
authors.forename: Chatchai
authors.initials: C
authors.lastname: Tayapiwatana
mesh_terms: D000595:Amino Acid Sequence; D017487:Ankyrins / Q000494:pharmacology*; D000998:Antiviral Agents / Q000494:pharmacology*; D036022:Capsid Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D018360:Crystallography, X-Ray; D004797:Enzyme-Linked Immunosorbent Assay; D015658:HIV Infections / Q000188:drug therapy / Q000821:virology; D015497:HIV-1 / Q000187:drug effects* / Q000254:growth & development; D006801:Humans; D008958:Models, Molecular; D008969:Molecular Sequence Data; D009154:Mutation / Q000235:genetics; D011485:Protein Binding; D011487:Protein Conformation; D025941:Protein Interaction Mapping; D017434:Protein Structure, Tertiary; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D017386:Sequence Homology, Amino Acid; D019065:Virus Assembly
chemical_list: D017487:Ankyrins; D000998:Antiviral Agents; D036022:Capsid Proteins; D011994:Recombinant Proteins