Abstract 1007 Novel Interactions of Potato Spindle Tuber Viroid RNA with Pokeweed Antiviral Protein

Potato spindle tuber viroid (PSTVd) (family Pospiviroidae) genome includes a small non-coding circular RNA molecule, and causes the spindle tuber disease in potatoes. Currently, successful control of PSTVd infection requires both prevention of infection and viroid eradication, necessitating destruction of the infected crops. Pokeweed antiviral protein (PAP) from Phytolacca americana has broad range of antiviral activities, selectively depurinating rRNA and viral RNAs, resulting in depressed viral infectivity. Structural similarities between the natural SRL substrate of PAP and the loop E of PSTVd genome allowed us to characterize these protein-RNA interactions quantitatively. We report a novel interaction between PAP and PSTVd RNA, and show that PAP binds to PSTVd RNA in a 1:1 stochiometric ratio, and this binding affinity elevates 3.6-fold as temperature decreases (from 25 °C to 5 °C). Formation of the PAP-PSTVd RNA complex is enthalpically-favored and entropically-driven. This results in overall negative binding energy (‒54.5 ± 1.3 kJ/mol). Our experimental observations are supported by the molecular modeling that places the conserved loop E of the PSTVd RNA into the RNA-binding pocket of PAP. Bioanalytical assay demonstrates that PAP depurinates viroid RNA, and kinetic findings show that over 90% of RNA depurination is complete after a 3-minute period. Altogether, the findings presented here are an initial study with promising results, providing a rational approach for controlling PSTVd infection. This work was supported by the John Jay College grant recovery fund and Seed Funding.