Structural determinants of HIV-1 nucleocapsid protein for cTAR DNA binding and destabilization, and correlation with inhibition of self-primed DNA synthesis.

The nucleocapsid protein (NC) of human immunodeficiency virus type 1 (HIV-1) is formed of two highly conserved CCHC zinc fingers flanked by small basic domains. NC is required for the two obligatory strand transfers in viral DNA synthesis through its nucleic acid chaperoning properties. The first DNA strand transfer relies on NC's ability to bind and destabilize the secondary structure of complementary transactivation response region (cTAR) DNA, to inhibit self-priming, and to promote the annealing of cTAR to TAR RNA.

8-vinyl-deoxyadenosine, an alternative fluorescent nucleoside analog to 2'-deoxyribosyl-2-aminopurine with improved properties.

We report here the synthesis and the spectroscopic characterization of 8-vinyl-deoxyadenosine (8vdA), a new fluorescent analog of deoxyadenosine. 8vdA was found to absorb and emit in the same wavelength range as 2'-deoxyribosyl-2-aminopurine (2AP), the most frequently used fluorescent nucleoside analog. Though the quantum yield of 8vdA is similar to that of 2AP, its molar absorption coefficient is about twice, enabling a more sensitive detection.

The C-terminal domain of the HIV-1 regulatory protein Vpr adopts an antiparallel dimeric structure in solution via its leucine-zipper-like domain.

HIV-1 Vpr is a highly conserved accessory protein that is involved in many functions of the virus life cycle. Vpr facilitates the entry of the HIV pre-integration complex through the nuclear pore, induces G2 cell cycle arrest, regulates cell apoptosis, increases transcription from the long terminal repeat and enhances viral replication. Vpr contains a Leu/Ile-rich domain (amino acids 60-81) in its C-terminal part, which is critical for dimerization.

Role of the structure of the top half of HIV-1 cTAR DNA on the nucleic acid destabilizing activity of the nucleocapsid protein NCp7.

The viral nucleic acid chaperone protein NCp7 of HIV-1 assists the two obligatory strand transfers required for the conversion of the genomic RNA into double-stranded DNA by reverse transcriptase. The first strand transfer necessitates the annealing of the early product of cDNA synthesis, the minus strand strong stop DNA (ss-cDNA) to the 3' end of the genomic RNA. The hybridization reaction involves regions containing imperfect stem-loop (SL) structures, namely the TAR RNA at the 3' end of the genomic RNA and the complementary sequence cTAR at the 3' end of ss-cDNA.

Impact of the terminal bulges of HIV-1 cTAR DNA on its stability and the destabilizing activity of the nucleocapsid protein NCp7.

Reverse transcription of HIV-1 genomic RNA to double-stranded DNA by reverse transcriptase (RT) is a critical step in HIV-1 replication. This process relies on two viral proteins, the RT enzyme and nucleocapsid protein NCp7 that has well documented nucleic acid chaperone properties. At the beginning of the linear DNA synthesis, the newly made minus-strand strong-stop DNA ((-)ssDNA) is transferred to the 3'end of the genomic RNA by means of an hybridization reaction between transactivation response element (TAR) RNA and cTAR DNA sequences.