Single aromatic residue location alters nucleic acid binding and chaperone function of FIV nucleocapsid protein

Hao Wu, Wei Wang, Nada Naiyer, Eric Fichtenbaum, Dominic F. Qualley, Micah J. McCauley, Robert J. Gorelick, Ioulia Rouzina, Karin Musier-Forsyth, Mark C. Williams

Research output: Contribution to journalArticlepeer-review

6 Scopus citations

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.

Original languageEnglish (US)
Pages (from-to)39-51
Number of pages13
JournalVirus research
Volume193
DOIs
StatePublished - Nov 26 2014

Bibliographical note

Funding Information:
This work was supported by the National Institutes of Health [ GM065056 to K.M.-F. and GM072462 to M.C.W.] and the National Science Foundation [ MCB-1243883 to M.C.W]. This project was also supported by the National Cancer Institute, National Institutes of Health [contract no. HHSN261200800001E with Leidos Biomedical Research, Inc. to R.J.G.]. The content of this publication does not necessarily reflect the views or policies of the Department of Health and Human Services, nor does mention of trade names, commercial products, or organizations imply endorsement by the U.S. Government. We thank Drs. Lawrence Mathes and Kathleen Hays (Ohio State University) for the DNA plasmid encoding FIV Gag. We also thank Donald G. Johnson and Catherine V. Hixson of the AIDS and Cancer Virus Program for their assistance in cloning, and producing and purifying several of the recombinant proteins used in this study.

Publisher Copyright:
© 2014 Elsevier B.V.

Copyright:
Copyright 2014 Elsevier B.V., All rights reserved.

Keywords

  • Aromatic residues
  • Feline immunodeficiency virus
  • Nucleic acid chaperone
  • Nucleocapsid protein
  • Single-molecule stretching
  • Zinc fingers

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