Poster abstracts
Poster number 30 submitted by Oliver Munyaradzi
Helical bPNA targetting U-rich RNA/T-rich DNA
Oliver Munyaradzi (Ohio State University), Dennis Bong (Ohio State University)
Abstract:
RNA-RNA and RNA-protein interactions are key players in aberrant pathways critical to many diseases. However, targeting such interactions presents a significant and largely unmet challenge. This project seeks to design RNA sequence-specific binders that take advantage of defined secondary structure. Previous work produced bifacial peptide nucleic acid (bPNA) in which melamine displayed on a modified lysine side-chain formed T-M-T/U-M-U base triples with thymine or uracil. We hypothesized that bPNA incorporating defined secondary structure would exhibit tighter binding, reducing the binding footprint and allowing us to target RNA-RNA and RNA-protein interactions.
Initially, we designed and synthesized two helical peptides by incorporating lysine whose side-chain was doubly melamine-functionalized into:
1) an i,i+4 alpha-helix repeat—Cbf-βAla-(AAAK2M)3-G
2) a poly-proline-II helix repeat—Cbf-βAla-(PPK2M)3-G
Secondary structure was probed by CD spectroscopy which confirmed that both peptides were helical. Biophysical properties were elucidated using Tm measurements and EMSA studies. For ssDNA T6C4T6 binding, no significant increase in Tm was noted (compared to previous bPNA) while 3-4 fold tighter binding was observed using EMSA. Surprisingly, the two helical peptides helped unstructured U6C4U6 ssRNA to fold, yielding complexes with a 10 oC higher Tm and with sufficient stability for EMSA studies compared to those formed by previous bPNA.
We anticipate that the helical peptides could be applied as reporters for RNA interactions and serve as means for detecting disease states.
Keywords: RNA-binding, helical peptide, protein-protein