1. The Wonderful World of RNA DNARNA protein

2. Complexity of RNA Folding 1 strand 1 strand 4 building blocks 4 building blocks Basic structural element: double helix Basic structural element: double helix RNA folds back and forms RNA folds back and forms Non-Watson Crick base pairs Non-Watson Crick base pairs base triples base triples Loop–loop, -single strand, -helix, Loop–loop, -single strand, -helix, helix - helix interactions helix - helix interactions The shape of an RNA molecule determines its function

3. RNA Base Pairs AU GC GUGA A G

4. I. RNA tetraloops Abundant in Nature Fullfill many structural and functional roles 16S rRNA:Of all loops > 50% are tetraloops Most are UNCG or GNRA Tetrahymena ribozyme

5. II. Cis-regulating elements of poliovirus Single stranded RNA genome 7440 nt

6. Poliovirus 5’-UTR Aim: To characterize the stem-loop D - 3CD interaction by using in vivo SELEX. Poliovirus 5´-UTR cloverleaf structure

7. N-NN-N N N-NN-N U-A NN N G-C random pool of 4 8 = 65536 possible sequences In vivo selection of randomly generated mutants N-NN-N N N-NN-N U-A NN N G-C 22 different selected sequences In vivo production of virus mutants A-U U U-G U-A GC G G-C polio wild type sequence

8. Structures of UNCG and GNRA loops differ Heus and Pardi, 1991 U G G C A U C G A A A G anti syn UG Du et al., 2003 anti GA

9. GCUA loop adopts a YNMG tetraloop configuration UACG loop GAAA loop GCUA loop C G A A A G UA U C A G U G G C A U

10. II. RNA pseudoknots Occur in virtually all classes of RNA Role in biological processes: - replication initiation - translational control - ribosomal frameshifting - core formation in larger RNAs

11. RNA pseudoknot formation

12. Ribosomal frameshifting SRV-1 genome overlapping region

13. NMR-derived structure of SRV-1 RNA pseudoknot

14. Guanine Riboswitch G OFF Guanine present Guanine absent ON Guanine

15. Structural Differences Flexible 5-residue “lid” Disordered base pairs in free aptamer Base pairs in free form and complex P1 arm P3 arm P2-P3 interaction in both the free form and complex P2 arm

16. Importance for riboswitching