Poster abstracts
Poster number 55 submitted by Madeline Glennon
Structural elucidation of an RNA triple helix in complex with a small molecule
Madeline M. Glennon (Department of Chemistry and Biochemistry, University of Notre Dame, Notre Dame, IN), Martina Zafferani (Department of Chemistry, Duke University, Durham, NC), Conner J. Langeberg (Innovative Genomics Institute, University of California, Berkeley, CA), Jeffery S. Kieft (New York Structural Biology Center, New York, NY), Amanda E. Hargrove (Department of Chemistry, Duke University, Durham, NC), Krishna M. Shivakumar, Jessica A. Brown (Department of Chemistry and Biochemistry, University of Notre Dame, Notre Dame, IN)
Abstract:
Human metastasis-associated lung adenocarcinoma transcript 1 (MALAT1) is a long non-coding RNA with a 3′-terminal triple helix, which stabilizes and protects the RNA from degradation. This stabilization contributes to MALAT1 overaccumulation, promoting cancer and disease. The unique structure and function of the MALAT1 triple helix makes it an ideal target for small molecule intervention. Yet, structural details regarding the interactions between the MALAT1 triple helix and a small molecule drug remain unclear. Herein, I aim to solve a 3D structure of the MALAT1 triple helix in complex with a diminazene (DMZ) small molecule: DMZp8. Single-particle cryo-electron microscopy (cryo-EM) is a technique most suitable for solving large macromolecular structures, yet can be applied to solve structures of small RNAs (<50 kDa). Visualization of small RNAs is limited by the signal-to-noise ratio, hindering global resolution. To overcome these limitations, RNA scaffolding techniques graft an RNA-of-interest onto a larger, well-structured RNA scaffold. Herein, we grafted the MALAT1 triple helix onto two established RNA scaffolds: TTR-3 (PDB ID: 6WLK) and a circularly permuted version of the Tetrahymena ribozyme (TetP6b) (PDB ID: 8TJX). Thus far, we have solved the unbound 3D structures of the MALAT1 triple helix-TTR-3 (5.39 Å) and the MALAT1 triple helix-dTetP6b (3.11 Å). Additionally, optimal single-particle density and distribution was observed for the MALAT1 triple helix-TTR-3:DMZp8 complex at a 1:4 ratio. Promising single-particle conditions for the MALAT1 triple helix-dTetP6b scaffold were achieved using a 1:25 ratio of the MALAT1 triple helix-dTetP6b:DMZp8. A high-resolution structure of a small molecule bound to the MALAT1 triple helix will advance the rational design of small molecules selective for disease-promoting RNAs.
Keywords: RNA triple helix, cryo-EM, small molecule