Poster abstracts
Poster number 112 submitted by Haoyun Yang
Dynamics of RNA Conformational Sampling by the SMK box riboswitch
Haoyun Yang (Department of chemistry and biochemistry, OSU), Mark Foster (Department of Chemistry and Biochemistry, OSU), Tina Henkin (Department of Microbiology, OSU)
Abstract:
The conformational dynamics of non-coding RNAs are coupled to vital cellular processes such as metabolite sensing, site specific RNA catalysis and the hierarchy of ordered ribonucleoprotein assemblies. However, the free energy landscapes that enable RNA functional conformational dynamics remain poorly understood. To fill this knowledge gap, we study a model system, the SMK box, S-adenosylmethionine (SAM) metabolite-binding/SAM-III riboswitch RNA, a compact RNA that undergoes ligand-mediated conformational changes to alter ribosome binding and thereby regulate translation1,2. Previously, X-ray crystallography analysis of the SMK aptamer bound to SAM established the structure of the ligand-bound, repressed state3. NMR, mutagenesis and chemical probing data suggest that in the absence of SAM, SMK is in equilibrium between an alternative fold (ISO) that has no obvious ligand binding site and a ligand binding-competent state (PRIMED)4,5. To understand how the SMK box riboswitch is able to undergo conformational fluctuations between alternative structures that enable its function we use stopped-flow measurements of SAM binding to 2-aminopurine labeled RNA to discriminate between conformational selection and induced-fit mechanisms of ligand binding. These experiments also quantify the thermodynamic barriers between conformations and provide structural insights into those rate-liming steps. Heteronuclear NMR spectroscopy is applied to determine the structures of the low-energy states, and the pathways connecting them.
References:
1. Fuchs RT, Grundy FJ, Henkin TM. S-adenosylmethionine directly inhibits binding of 30S ribosomal subunits to the SMK box translational riboswitch RNA. Proc Natl Acad Sci U S A. 2007;104(12)
2. Fuchs RT, Grundy FJ, Henkin TM. The S(MK) box is a new SAM-binding RNA for translational regulation of SAM synthetase. Nat Struct Mol Biol. 2006;13(3)
3. Lu C, Smith AM, Fuchs RT, et al. Crystal structures of the SAM-III/S(MK) riboswitch reveal the SAM-dependent translation inhibition mechanism. Nat Struct Mol Biol. 2008;15(10)
4. Lu C, Smith AM, Ding F, Chowdhury A, Henkin TM, Ke A. Variable sequences outside the SAM-binding core critically influence the conformational dynamics of the SAM-III/SMK box riboswitch. J Mol Biol. 2011;409(5)
5. Wilson RC, Smith AM, Fuchs RT, Kleckner IR, Henkin TM, Foster MP. Tuning riboswitch regulation through conformational selection. J Mol Biol. 2011;405(4)
Keywords: Riboswitch , Dynamics , NMR