Validation of a novel secretion modification region (SMR) of HIV-1 Nef using cohort sequence analysis and molecular modeling
- 278 Downloads
The HIV-1 accessory protein Nef plays an active role in the pathogenesis of AIDS by its numerous cellular interactions that facilitate the release of virus particles. This 27 kDa protein is required for maintenance of the viral replication in HIV, and is also known to contribute to immune evasion, blocking of apoptosis in virus-infected cells and enhancement of virus infectivity. Nef has been shown to be secreted and is present on the surface of virus-infected cells. Recent studies from our laboratory have shown that the Nef protein is secreted from nef-transfected and HIV-1-infected cells in small exosome-like vesicles (40–100 nm diam.) that do not contain virions. We have identified three amino-terminal domains of Nef as necessary for secretion: (i) the four arginine residues (17,19,21, 22) comprising the basic region; (ii) the phosphofurin acidic cluster sequence (PACS) composed of four glutamic acid residues (61–64); (iii) a previously unknown motif spanning amino acid residues 65–69 (VGFPV) which we named the secretion modification region (SMR). In this study, we have used population-based phylogeny data and sequence analysis to characterize the conservation of the Nef SMR domain that regulates vesicle secretion. We have performed in silico computational chemistry analysis involving molecular dynamic structure modeling of mutations in the SMR motif. Sequence analysis of Nef from HIV-1-infected patients, including slow progressors (SP), long term progressors (LTP) and long term non-progressors (LTNP) demonstrated 99 % conservation of the Nef SMR motif. Computational analysis including modeling of wild-type HIV-1 Nef and V66A Nef SMR mutant using structural homology and molecular dynamics of ligand-associated interactions indicated significant structural changes in the Nef mutant, thus supporting the importance of the SMR domain for mediating Nef vesicle secretion.
KeywordsHIV-1 protein Homology modeling Molecular dynamics Structure-function relationship
This work and the researchers at Morehouse were supported by NIH/NIGMS/MBRS grant S06 GM08248, NIH/NCRR/RCMI grant G12-RR03034, Georgia Research Alliance funding grant GRA.VAC08.W. This investigation was conducted in a facility constructed with support from Research Facilities Improvement Grant #C06 RR18386 from NIH/NCRR. The work, researchers, and facilities at Jackson State University NSF CREST Interdisciplinary Nanotoxicity Center were supported by grant NSF-CREST - # HRD-0833178. No competing financial interests exist.
- 4.Priceputu E, Rodrigue I, Chrobak P, Poudrier J, Mak TW, Hanna Z, Hu C, Kay DG, Jolicoeur P (2005) The Nef-mediated AIDS-like disease of CD4C/human immunodeficiency virus transgenic mice is associated with increased Fas/FasL expression on T cells and T-cell death but is not prevented in Fas-, FasL-, tumor necrosis factor receptor 1-, or interleukin-1beta-converting enzyme-deficient or Bcl2-expressing transgenic mice. J Virol 79:6377–6391CrossRefGoogle Scholar
- 14.Zeigler ZR, Rosenfeld CS, Andrews DF III, Nemunaitis J, Raymond JM, Shadduck RK, Kramer RE, Gryn JF, Rintels PB, Besa EC, George JN (1996) Plasma von Willebrand Factor Antigen (vWF:AG) and thrombomodulin (TM) levels in Adult Thrombotic Thrombocytopenic Purpura/Hemolytic Uremic Syndromes (TTP/HUS) and bone marrow transplant-associated thrombotic microangiopathy (BMT-TM). Am J Hematol 53:213–220CrossRefGoogle Scholar
- 20.Percario Z, Olivetta E, Fiorucci G, Mangino G, Peretti S, Romeo G, Affabris E, Federico M (2003) Human immunodeficiency virus type 1 (HIV-1) Nef activates STAT3 in primary human monocyte/macrophages through the release of soluble factors: involvement of Nef domains interacting with the cell endocytotic machinery. J Leukocyte Biol 74:821–832CrossRefGoogle Scholar
- 21.Campbell TD, Khan M, Huang M-B, Bond VC, Powell MD (2008) HIV-1 Nef protein is secreted into vesicles that can fuse with target cells and virions. Ethn Dis 18:S2–S9Google Scholar
- 23.Leitner T, Foley B, Hahn B, Marx P, McCutchan F, Mellors J, Wolinsky S, Korber B, (2007) HIV Sequence Compendium 2006/2007. Los Alamos National Laboratory, NM 07–Google Scholar
- 34.Petsko GA, Dagmar R (2003) Protein structure and function. Wiley-Blackwell, LondonGoogle Scholar