title: RNA recognition mechanism of the minimal active domain of the human immunodeficiency virus type-2 nucleocapsid protein.
abstract: NCp8 of HIV-2 contains two CCHC-type zinc fingers connected by a linker, and is involved in many critical steps of the virus life cycle. It was previously shown that the first zinc finger flanked by the linker is the minimal active domain for specific binding to viral RNA. In our previous study, we determined the three-dimensional structure of NCp8-f1, including the minimal active domain, and found that a hydrogen bond between Asn(11) N(delta)H and Arg(27) O stabilized the conformation of the linker in the vicinity of the zinc finger [Kodera et al. (1998) Biochemistry 37, 17704-17713]. In this study, RNA binding activities of NCp8-f1 and three types of its mutant peptides were analysed by native PAGE assay. The activity and three-dimensional structure of NCp8-f1/N11A, in which alanine is substituted for Asn(11) thereby affecting the conformation of the linker, was analyzed and compared with those of NCp8-f1. We demonstrated that the existence of Arg(4) and/or Lys(5) and Arg(26) and/or Arg(27) were necessary for binding RNA. Furthermore, the linker's flexible orientation, which is controlled by the hydrogen bond between Asn(11) N(delta)H and Arg(27) O, appears to be a structural basis for NCp8 existing as a multi-functional protein.
authors.affiliation: Department of Physics, School of Science, Kitasato University, Sagamihara, Kanagawa 228-8555, Japan.
authors.forename: Takashi
authors.initials: T
authors.lastname: Matsui
authors.forename: Yoshio
authors.initials: Y
authors.lastname: Kodera
authors.forename: Hiroshi
authors.initials: H
authors.lastname: Endoh
authors.forename: Emi
authors.initials: E
authors.lastname: Miyauchi
authors.forename: Hiroyoshi
authors.initials: H
authors.lastname: Komatsu
authors.forename: Kazuki
authors.initials: K
authors.lastname: Sato
authors.forename: Takeshi
authors.initials: T
authors.lastname: Tanaka
authors.forename: Toshiyuki
authors.initials: T
authors.lastname: Kohno
authors.forename: Tadakazu
authors.initials: T
authors.lastname: Maeda
mesh_terms: D036022:Capsid Proteins / Q000737:chemistry* / Q000378:metabolism*; D015498:HIV-2 / Q000737:chemistry*; D006801:Humans; D009154:Mutation; D010455:Peptides; D017434:Protein Structure, Tertiary; D012367:RNA, Viral / Q000737:chemistry / Q000378:metabolism*; D016601:RNA-Binding Proteins / Q000737:chemistry* / Q000378:metabolism*; D016335:Zinc Fingers
chemical_list: D036022:Capsid Proteins; D010455:Peptides; D012367:RNA, Viral; D016601:RNA-Binding Proteins; C101406:nucleocapsid protein 8, HIV-2