The nucleocapsid protein (NC) of HIV-1 is a 55 amino acid long protein containing two zinc fingers and many basic residues, especially on the unstructured N-terminal and linker region between the two fingers. NC binds nucleic acids with high affinity and condenses double-stranded (ds) DNA into compact structures. While condensation is mediated by basic residues of NC, consistent with counter ion condensation theory, NC is a more efficient condensing agent than other cationic polyamines such as spermine, protamine, and poly-arginine. We utilize optical tweezers, confocal imaging, and atomic force microscopy to measure and manipulate NC-dsDNA condensates, which spontaneously form on long dsDNA templates at sufficient NC concentration or protein-nucleotide ratio. These compact structures stabilize the NC-dsDNA interaction, causing sequestration of NC to the condensed dsDNA complex relative to the sparsely bound, uncondensed dsDNA regions. While individual NC proteins dissociate from dsDNA on a ten second timescale, the NC-dsDNA condensate, once formed, resists disassembly for tens of minutes even in the absence of free protein. During the viral replication cycle, completion of reverse transcription results in the HIV-1 capsid containing a full-length dsDNA proviral genome (∼10 kb) and a similar quantity of RNA template, which was original packaged in the viral particle, along with over one thousand NC proteins. Our in vitro studies suggest NC-dsDNA globules formed in the capsid slowly disentangle, likely delaying intra-capsid pressure build up and subsequent uncoating. Additionally, this globule likely does not dissipate immediately when the capsid structure begins disassembling, and thus NC may still be present on the proviral dsDNA during later stages of infection. This potentially explains why particular NC mutations have been found to affect the timing of uncoating and/or integration, conferring partial immunity to certain HIV inhibitors.
Morse et al. (Sun,) studied this question.