Mutational Analysis of Two Zinc Finger Motifs in HIV Type 1 Nucleocapsid Proteins: Effects on Proteolytic Processing of Gag Precursors and Particle Formation

The NCps (nucleocapsid proteins) of HIV-1 (HIV type 1), HIV-2 and SIV (simian immunodeficiency virus) are small highly basic proteins, characterized by the presence of two CCHC ZF (zinc finger) domains. NCps, closely associated with the dimeric RNA genome in the core of the virus particle, were shown to promote the specific encapsidation of the viral RNA and are implicated in reverse transcription. Solution structure of the HIV-1 NCp7 and complexes of NCp7 with RNA or DNA showed the critical relationships between the structure and its various functions. HIV-1 and HIV-2 have resulted respectively from transmissions of SIV from chimpanzees and sooty mangabeys. It has been shown that the SIVlhoest (SIV from l'Hoest monkeys) also has the potential to infect human populations. Since monkeys are of great interest for clinical studies of antiviral drugs, the structure of (13-51)NCp8 (zinc finger domain of NCp8, encompassing residues 13–51) from SIVlhoest was determined by NMR to appraise the influence of major differences in the sequence, since Glu21, Gly43 and Met46 in NCp7 are replaced by Pro, Glu and Phe respectively in this particular NCp8. The structure of (13-51)NCp8 is very well defined, and surprisingly the structure of each ZF is similar in NCp7 and NCp8. Moreover, contrary to NCp7, the two ZFs are strongly locked to each other in this NCp8. This first reported structure of a simian NCp8 compared with that of NCp7 shows that the main structural differences occur at the flexible linker between the two ZFs but the essential residues responsible for the interaction with oligonucleotides adopt the same orientation in the two proteins.

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Effects of HIV Type 1 Envelope Glycoprotein Proteolytic Processing on Antigenicity

AIDS Research and Human Retroviruses ◽

10.1089/088922203763315722 ◽

2003 ◽

Vol 19 (3) ◽

pp. 217-226 ◽

Cited By ~ 43

Author(s):

Zhihai Si ◽

Ngoc Phan ◽

Enko Kiprilov ◽

Joseph Sodroski

Keyword(s):

Envelope Glycoprotein ◽

Proteolytic Processing ◽

Hiv Type 1

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Wild-Type and Mutant HIV Type 1 Nucleocapsid Proteins Increase the Proportion of Long cDNA Transcripts by Viral Reverse Transcriptase

AIDS Research and Human Retroviruses ◽

10.1089/aid.1997.13.533 ◽

1997 ◽

Vol 13 (7) ◽

pp. 533-543 ◽

Cited By ~ 29

Author(s):

JAMES E. DRUMMOND ◽

PHOEBE MOUNTS ◽

ROBERT J. GORELICK ◽

JOSE R. CASAS-FINET ◽

WILLIAM J. BOSCHE ◽

...

Keyword(s):

Reverse Transcriptase ◽

Wild Type ◽

Nucleocapsid Proteins ◽

Hiv Type 1

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Interaction with the p6 Domain of the Gag Precursor Mediates Incorporation into Virions of Vpr and Vpx Proteins from Primate Lentiviruses

Journal of Virology ◽

10.1128/jvi.73.1.592-600.1999 ◽

1999 ◽

Vol 73 (1) ◽

pp. 592-600 ◽

Cited By ~ 89

Author(s):

L. Selig ◽

J.-C. Pages ◽

V. Tanchou ◽

S. Prévéral ◽

C. Berlioz-Torrent ◽

...

Keyword(s):

Nucleocapsid Protein ◽

Proteolytic Processing ◽

Triplet Repeat ◽

Repeat Region ◽

Gag Proteins ◽

Primate Lentiviruses ◽

Hiv Type 1 ◽

Equivalent Region ◽

Hiv 1

ABSTRACT Vpr and Vpx proteins from human and simian immunodeficiency viruses (HIV and SIV) are incorporated into virions in quantities equivalent to those of the viral Gag proteins. We demonstrate here that Vpr and Vpx proteins from distinct lineages of primate lentiviruses were able to bind to their respective Gag precursors. The capacity of HIV type 1 (HIV-1) Vpr mutants to bind to Pr55Gag was correlated with their incorporation into virions. Molecular analysis of these interactions revealed that they required the C-terminal p6 domain of the Gag precursors. While the signal for HIV-1 Vpr binding lies in the leucine triplet repeat region of the p6 domain reported to be essential for incorporation, SIVsm Gag lacking the equivalent region still bound to SIVsm Vpr and Vpx, indicating that the determinants for Gag binding are located upstream of this region of the p6 domain. Binding to Gag cleavage products showed that HIV-1 Vpr interacted directly with the nucleocapsid protein (NC), whereas SIVsm Vpr and Vpx did not interact with NC but with the p6 protein. These results (i) reveal differences between HIV-1 and SIVsm for the p6 determinants required for Vpr and Vpx binding to Gag and (ii) suggest that HIV-1 Vpr and SIVsmVpr and Vpx interact with distinct cleavage products of the precursor following proteolytic processing in the virions.

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Mutational analysis of two zinc-finger motifs in the nucleocapsid protein of simian immunodeficiency virus mac239

Journal of General Virology ◽

10.1099/vir.0.18865-0 ◽

2003 ◽

Vol 84 (6) ◽

pp. 1641-1648 ◽

Cited By ~ 7

Author(s):

Wataru Akahata ◽

Eiji Ido ◽

Masanori Hayami

Keyword(s):

Simian Immunodeficiency Virus ◽

Zinc Finger ◽

Nucleocapsid Protein ◽

Mutational Analysis ◽

Zinc Finger Motifs ◽

Immunodeficiency Virus

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Generation of Monoclonal Antibodies Specifically Directed against the Proximal Zinc Finger of HIV Type 1 NCp7

AIDS Research and Human Retroviruses ◽

10.1089/08892220050117023 ◽

2000 ◽

Vol 16 (13) ◽

pp. 1259-1267 ◽

Cited By ~ 4

Author(s):

Hugues De Rocquigny ◽

Anne Caneparo ◽

Chang-Zhi Dong ◽

Thierry Delaunay ◽

Bernard P. Roques

Keyword(s):

Monoclonal Antibodies ◽

Zinc Finger ◽

Hiv Type 1

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Resistance Mutational Analysis of HIV Type 1 Subtype C among Rural South African Drug-Naive Patients Prior to Large-Scale Availability of Antiretrovirals

AIDS Research and Human Retroviruses ◽

10.1089/aid.2006.22.1306 ◽

2006 ◽

Vol 22 (12) ◽

pp. 1306-1312 ◽

Cited By ~ 20

Author(s):

Pascal O. Bessong ◽

Jeffrey Mphahlele ◽

Isaac A. Choge ◽

Larry C. Obi ◽

Lynn Morris ◽

...

Keyword(s):

South African ◽

Large Scale ◽

Mutational Analysis ◽

Subtype C ◽

Rural South ◽

Drug Naïve ◽

Hiv Type 1

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Importance of Basic Residues in the Nucleocapsid Sequence for Retrovirus Gag Assembly and Complementation Rescue

Journal of Virology ◽

10.1128/jvi.72.11.9034-9044.1998 ◽

1998 ◽

Vol 72 (11) ◽

pp. 9034-9044 ◽

Cited By ~ 92

Author(s):

J. Bradford Bowzard ◽

Robert P. Bennett ◽

Neel K. Krishna ◽

Sandra M. Ernst ◽

Alan Rein ◽

...

Keyword(s):

Zinc Finger ◽

Mutational Analysis ◽

Zinc Fingers ◽

Leukemia Virus ◽

Foamy Virus ◽

Gag Proteins ◽

Amino Terminal ◽

C Terminus ◽

Zinc Finger Motifs ◽

And Function

ABSTRACT The Gag proteins of Rous sarcoma virus (RSV) and human immunodeficiency virus (HIV) contain small interaction (I) domains within their nucleocapsid (NC) sequences. These overlap the zinc finger motifs and function to provide the proper density to viral particles. There are two zinc fingers and at least two I domains within these Gag proteins. To more thoroughly characterize the important sequence features and properties of I domains, we analyzed Gag proteins that contain one or no zinc finger motifs. Chimeric proteins containing the amino-terminal half of RSV Gag and various portions of the carboxy terminus of murine leukemia virus (MLV) (containing one zinc finger) Gag had only one I domain, whereas similar chimeras with human foamy virus (HFV) (containing no zinc fingers) Gag had at least two. Mutational analysis of the MLV NC sequence and inspection of I domain sequences within the zinc-fingerless C terminus of HFV Gag suggested that clusters of basic residues, but not the zinc finger motif residues themselves, are required for the formation of particles of proper density. In support of this, a simple string of strongly basic residues was found to be able to substitute for the RSV I domains. We also explored the possibility that differences in I domains (e.g., their number) account for differences in the ability of Gag proteins to be rescued into particles when they are unable to bind to membranes. Previously published experiments have shown that such membrane-binding mutants of RSV and HIV (two I domains) can be rescued but that those of MLV (one I domain) cannot. Complementation rescue experiments with RSV-MLV chimeras now map this difference to the NC sequence of MLV. Importantly, the same RSV-MLV chimeras could be rescued by complementation when the block to budding was after, rather than before, transport to the membrane. These results suggest that MLV Gag molecules begin to interact at a much later time after synthesis than those of RSV and HIV.

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