1. eE-Motif Structures in DNA Kathryn Iverson USC Health Sciences Center and School of Pharmacy SoCalBSI 2007

2. Outline E-motif structure Project objectives Methods Working results

3. 5’ CCGCCGCCG CGCCGCCG CCGCCGCC G C C GCCGCCG GCGGCGGCGGCGGCGG CGGCGGCGGCGGCGGC G GC G CC G C GC GC CG CC C G G G G G C C G G C G Hairpins formed by GCC repeats

5. 5’ C GC CG GC CG C 5’ C C

6. E-motif Structure Extra-helical cytosines Extra-helical cytosines from first GCC and third GCC repeats are interacting Stacking between G-C

7. Fragile X Is the most common forms of inherited mental retardation. 1 in 2000 boys and 1 in 4000 girls are afflicted and 1 in 260 women are carriers 1 July 22, National Fragile X Awareness Day Characterized by the expansion of GCC repeat in the 5’ untranslated region of the FMR1 gene 1. http://www.conquerfragilex.org/about.php

8. Objectives of Project Accurate model of eE-motif structure Knowledge of extra-helical cytosine structures Insight into Fragile X syndrome Therapeutic targets

9. Methods Generate NASDAC input files Run energy minimization on the structures using AMBER Molecular dynamics modeling with restraints in AMBER

10. Generation of pdb files User enters how many GCC repeats Option to protonate one of cytosines What parameter to vary and within what range Create data.in file, run NASDAC and produce.pdb file

11. Minimization Without Restraints G-C base pairs no longer close enough to hydrogen bond Stacking of extra- helical cytosines

12. Restraints Hydrogen bonding distance between G and C base pairs Distance between N2 atoms in G-C base pairs Torsion angle for G-C base pairs Distance between extra-helical cytosines

13. eE-Motif with Restraints Extra-helical cytosines are in the same plane Hydrogen bonding

14. Acknowledgements Thank you to Dr. Ian Haworth and Dr. Rebecca Romero for their guidance and advice throughout the project USC Health Sciences Center and School of Pharmacy Program leaders and faculty of SoCalBSI SoCalBSI participants Cal State LA

15. Questions?