Nucleic Acid Chaperone Activity of Wild Type and Mutant FIV Nucleocapsid Proteins

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The feline immunodeficiency virus (FIV) nucleocapsid (NC) protein contains 66 amino acids. It has similar composition to HIV-1 NC, with a high number of positively charged amino acids and two zinc fingers, each having a single aromatic residue (F12 and W44). However, there are some differences between the structures of these two proteins, which likely contribute to their different nucleic acid chaperone properties. In addition to a short N-terminal basic domain, FIV NC has a C-terminal positively charged tail. Moreover, whereas HIV-1 NC’s two aromatic residues are located on the same side of each zinc finger domain, FIV NC’s aromatic residues are located on opposite sides of the zinc fingers. It is known that HIV-1 NC has optimal chaperone activity, including the ability to strongly aggregate nucleic acids, destabilize nucleic acid secondary structure and facilitate rapid protein-nucleic acid interaction kinetics. We use single molecule experiments to measure the characteristics of wild type and mutant FIV NC. By stretching single DNA molecules in the presence of these proteins, we measure their ability to induce aggregation, stabilize or destabilize dsDNA, and facilitate nucleic acid annealing. Our results show that wild type FIV NC induces significant DNA aggregation, but instead of destabilizing double-stranded DNA, it appears to stabilize the DNA. FIV NC variants containing mutations in the aromatic residues, basic residues, and zinc finger residues possess strongly altered nucleic acid chaperone properties. This work allows us to directly relate FIV NC structure with its function. The single molecule experiments are also compared to ensemble nucleic acid binding and chaperone studies. Taken together, this study sheds new light on the mechanism by which specific types and locations of residues contribute to NC’s nucleic acid chaperone activity and retroviral replication in these systems.