Specific Nucleic Acid Chaperone Activity of HIV-1 Nucleocapsid Protein Deduced from Hairpin Unfolding
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RNA and DNA hairpin formation and disruption play key regulatory roles in a variety of cellular processes. The 59-nucleotide transactivation response (TAR) RNA hairpin facilitates the production of full-length transcripts of the HIV-1 genome. Yet the stability of this long, irregular hairpin becomes a liability during reverse transcription as 24 base pairs must be disrupted for strand transfer. Retroviral nucleocapsid (NC) proteins serve as nucleic acid chaperones that have been shown to both destabilize the TAR hairpin and facilitate strand annealing with its complementary DNA sequence. Yet it has remained difficult to elucidate the way NC targets and dramatically destabilizes this hairpin while only weakly affecting the annealed product. In this work, we used optical tweezers to measure the stability of TAR and found that adding NC destabilized the hairpin and simultaneously caused a distinct change in both the height and location of the energy barrier. This data was matched to an energy landscape predicted from a simple theory of definite base pair destabilization. Comparisons revealed the specific binding sites found by NC along the irregular TAR hairpin. Furthermore, specific binding explained both the unusual shift in the transition state and the much weaker effect on the annealed product. These experiments illustrate a general method of energy landscape transformation that exposes important physical insights.
Key wordsEnergy landscape Transition state Nucleic acid hairpin Reverse transcription Optical tweezers Nucleic acid-protein interactions Mfold
This work was supported by grants NIH GM072462 and NSF MCB-1817712 to M.C.W.
- 47.Crooks GE (1999) Entropy production fluctuation theorem and the nonequilibrium work relation for free energy differences. Phys Rev E 60(3):2721–2726Google Scholar
- 48.Bennett CH (1976) Efficient estimation of free energy differences from Monte Carlo data. J Comp Phys 22(2):245–268Google Scholar