Talk Abstracts

Talk on Friday 04:06-04:18pm submitted by Daniel Binzel

Entropy As a Key Factor for One Step Formation of Phi29 pRNA 3WJ from Three RNA Fragments

Daniel Binzel (Nanobiotechnology Center, Markey Cancer Center, College of Pharmacy, University of Kentucky, Lexington, KY 40536, USA), Emil Khisamutdinov (Nanobiotechnology Center, Markey Cancer Center, College of Pharmacy, University of Kentucky, Lexington, KY 40536, USA), Peixuan Guo (Nanobiotechnology Center, Markey Cancer Center, College of Pharmacy, University of Kentucky, Lexington, KY 40536, USA)

Abstract:
The applicational use of functional RNA nanoparticles requires the creation of chemically and thermodynamically stable RNA core scaffolds. We have discovered an ultra-stable three-way junction (3WJ) in the bacteriophage phi29 motor packaging RNA (pRNA). This 3WJ can assemble from three short RNA oligos with high efficiency. Thermodynamic studies during assembly of the 3WJ indicated no dimer intermediate formation as the RNAs were only seen as monomer components or assembled three way junction; meaning the 3WJ does not follow the usually seen single assembly pathway of A + B + C ↔ AB + C ↔ ABC. A + B + C leads to the formation of ABC efficiently within a rapid time. Additionally, the 3WJ displayed high thermodynamic stability by exhibiting a melting temperature (T_M) of 59.3 ± 1.7 °C with the substitution of 2’-fluoro (2’-F) nucleotides increasing the T_M to 69.8 ± 2.0 °C, while the DNA 3WJ displayed a melting temperature of only 48.9 ± 3.2 °C. Furthermore, the Gibbs free energy of formations (ΔG°₃₇) were found to be -36, -28, and -15 kcal/mol for 3WJ_2’-F, 3WJ_RNA, and 3WJ_DNA, respectively. Surprisingly, the added stability, indicated by the more negative ΔG°₃₇, in the two RNA species of the pRNA-3WJ was attributed to the more favorable change in entropy (ΔS°), compared to DNA instead of the commonly seen more favorable enthalpy change. The unique thermodynamic stability and folding pathway of the pRNA-3WJ makes it an ideal candidate for RNA nanoparticles and nanoscaffolds.

References:
1. Guo P. The emerging field of RNA nanotechnology. Nature Nanotechnology. 2010; 5(12):833-42.

2. Shu D, Shu Y, Haque F, Abdelmawla S, Guo P. Thermodynamically stable RNA three-way junction as a platform for constructing multifunctional nanoparticles for delivery of therapeutics. Nature Nanotechnology. 2011; 6(10):658-67.

3. Binzel DW, Khisamutdinov EF, Guo P. Entropy-Driven One-Step Formation of Phi29 pRNA 3WJ from Three RNA Fragments. Biochemistry. 2014 Apr 15; 53(14):2221-31.

Keywords: Phi29, Thermodynamics, RNA Nanotechnology