title: Single aromatic residue location alters nucleic acid binding and chaperone function of FIV nucleocapsid protein.
abstract: Feline immunodeficiency virus (FIV) is a retrovirus that infects domestic cats, and is an excellent animal model for human immunodeficiency virus type 1 (HIV-1) pathogenesis. The nucleocapsid (NC) protein is critical for replication in both retroviruses. FIV NC has several structural features that differ from HIV-1 NC. While both NC proteins have a single conserved aromatic residue in each of the two zinc fingers, the aromatic residue on the second finger of FIV NC is located on the opposite C-terminal side relative to its location in HIV-1 NC. In addition, whereas HIV-1 NC has a highly charged cationic N-terminal tail and a relatively short C-terminal extension, the opposite is true for FIV NC. To probe the impact of these differences on the nucleic acid (NA) binding and chaperone properties of FIV NC, we carried out ensemble and single-molecule assays with wild-type (WT) and mutant proteins. The ensemble studies show that FIV NC binding to DNA is strongly electrostatic, with a higher effective charge than that observed for HIV-1 NC. The C-terminal basic domain contributes significantly to the NA binding capability of FIV NC. In addition, the non-electrostatic component of DNA binding is much weaker for FIV NC than for HIV-1 NC. Mutation of both aromatic residues in the zinc fingers to Ala (F12A/W44A) further increases the effective charge of FIV NC and reduces its non-electrostatic binding affinity. Interestingly, switching the location of the C-terminal aromatic residue to mimic the HIV-1 NC sequence (N31W/W44A) reduces the effective charge of FIV NC and increases its non-electrostatic binding affinity to values similar to HIV-1 NC. Consistent with the results of these ensemble studies, single-molecule DNA stretching studies show that while WT FIV NC has reduced stacking capability relative to HIV-1 NC, the aromatic switch mutant recovers the ability to intercalate between the DNA bases. Our results demonstrate that altering the position of a single aromatic residue switches the binding mode of FIV NC from primarily electrostatic binding to more non-electrostatic binding, conferring upon it NA interaction properties comparable to that of HIV-1 NC.
authors.affiliation: Northeastern University, Department of Physics, Boston, MA 02115, USA.
authors.forename: Hao
authors.initials: H
authors.lastname: Wu
authors.affiliation: The Ohio State University, Department of Chemistry and Biochemistry, Center for Retrovirus Research, and Center for RNA Biology, Columbus, OH 43210, USA.
authors.forename: Wei
authors.initials: W
authors.lastname: Wang
authors.affiliation: The Ohio State University, Department of Chemistry and Biochemistry, Center for Retrovirus Research, and Center for RNA Biology, Columbus, OH 43210, USA.
authors.forename: Nada
authors.initials: N
authors.lastname: Naiyer
authors.affiliation: The Ohio State University, Department of Chemistry and Biochemistry, Center for Retrovirus Research, and Center for RNA Biology, Columbus, OH 43210, USA.
authors.forename: Eric
authors.initials: E
authors.lastname: Fichtenbaum
authors.affiliation: The Ohio State University, Department of Chemistry and Biochemistry, Center for Retrovirus Research, and Center for RNA Biology, Columbus, OH 43210, USA.
authors.forename: Dominic F
authors.initials: DF
authors.lastname: Qualley
authors.affiliation: Northeastern University, Department of Physics, Boston, MA 02115, USA.
authors.forename: Micah J
authors.initials: MJ
authors.lastname: McCauley
authors.affiliation: AIDS and Cancer Virus Program, Leidos Biomedical Research, Inc., Frederick National Laboratory for Cancer Research, Frederick, MD 21702, USA.
authors.forename: Robert J
authors.initials: RJ
authors.lastname: Gorelick
authors.affiliation: University of Minnesota, Department of Biochemistry, Molecular Biology, and Biophysics, Minneapolis, MN 55455, USA.
authors.forename: Ioulia
authors.initials: I
authors.lastname: Rouzina
authors.affiliation: The Ohio State University, Department of Chemistry and Biochemistry, Center for Retrovirus Research, and Center for RNA Biology, Columbus, OH 43210, USA. Electronic address: musier@chemistry.ohio-state.edu.
authors.forename: Karin
authors.initials: K
authors.lastname: Musier-Forsyth
authors.affiliation: Northeastern University, Department of Physics, Boston, MA 02115, USA. Electronic address: mark@neu.edu.
authors.forename: Mark C
authors.initials: MC
authors.lastname: Williams
mesh_terms: D000595:Amino Acid Sequence; D019943:Amino Acid Substitution; D000818:Animals; D020029:Base Pairing; D002352:Carrier Proteins; D002415:Cats; D016089:Immunodeficiency Virus, Feline / Q000235:genetics* / Q000378:metabolism*; D035683:MicroRNAs / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D018832:Molecular Chaperones / Q000378:metabolism; D008969:Molecular Sequence Data; D009154:Mutation; D009690:Nucleic Acid Conformation; D019590:Nucleocapsid Proteins / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism*; D011485:Protein Binding; D012367:RNA, Viral / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism*
chemical_list: D002352:Carrier Proteins; D035683:MicroRNAs; D018832:Molecular Chaperones; D019590:Nucleocapsid Proteins; D012367:RNA, Viral
keywords: Aromatic residues; Feline immunodeficiency virus; Nucleic acid chaperone; Nucleocapsid protein; Single-molecule stretching; Zinc fingers