Title | An intrabody based on a llama single-domain antibody targeting the N-terminal alpha-helical multimerization domain of HIV-1 rev prevents viral production. |
Publication Type | Journal Article |
Year of Publication | 2010 |
Authors | Vercruysse, T., E. Pardon, E. Vanstreels, J. Steyaert, and D. Daelemans |
Journal | J Biol Chem |
Volume | 285 |
Issue | 28 |
Pagination | 21768-80 |
Date Published | 2010 Jul 9 |
ISSN | 1083-351X |
Keywords | Active Transport, Cell Nucleus, Amino Acid Sequence, Animals, Anti-HIV Agents, Antibodies, Camelids, New World, Dimerization, Fluorescence Resonance Energy Transfer, Gene Products, rev, HeLa Cells, HIV Infections, HIV-1, Humans, Lysine, Molecular Sequence Data, Protein Interaction Mapping, Protein Structure, Secondary, Protein Structure, Tertiary, Tyrosine |
Abstract | The human immunodeficiency virus, type 1 (HIV-1)-encoded Rev protein is essential for the expression of late viral mRNAs. Rev forms a large organized multimeric protein-protein complex on the Rev response element of these viral mRNA species and transports them from the nucleus to the cytoplasm, exploiting the CRM1-mediated cellular machinery. Here we report the selection of a nanobody, derived from a llama heavy-chain only antibody, that efficiently blocks the assembly of Rev multimers. The nanobody inhibits HIV-1 replication in cells and specifically suppresses the Rev-dependent expression of partially spliced and unspliced HIV-1 RNA. In HIV-susceptible cells, this nanobody thus has potential as an effective anti-HIV agent using genetic immunization strategies. Its binding site was mapped to Rev residues Lys-20 and Tyr-23 located in the N-terminal alpha-helical multimerization domain. In the presence of this nanobody, we observed an accumulation of dimeric Rev species, supporting a head-to-head/tail-to-tail molecular model for Rev assembly. The results indicate that the oligomeric assembly of Rev follows an ordered stepwise process and identify a new epitope within Rev that could guide strategies for the development of novel HIV inhibitors. |
DOI | 10.1074/jbc.M110.112490 |
Alternate Journal | J. Biol. Chem. |
PubMed ID | 20406803 |
PubMed Central ID | PMC2898381 |
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