Poster abstracts
Poster number 46 submitted by Haoyun Yang
Investigation of the dynamics of 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 detailed mechanistic description of the structures and RNA functional conformational dynamics remain poorly understood. To fill this knowledge gap, we propose to 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. In the absence of SAM, SMK is proposed to be in equilibrium between an alternative fold (ISO) that has no obvious ligand binding site and a ligand binding competent state (PRIMED) evidenced by mutation and chemical probing experiments4,5. Thus, a conformation selection model is speculated. To understand how the SMK box riboswitch is able to undergo conformational fluctuations between alternative structures in order to enable its function. We propose to first use stopped-flow and 2-aminopurine labeled RNA to reveal the ligand binding model that best describes this riboswitch and extract thermodynamics parameters that can provide structural insight of the binding steps. From this, we can use NMR relaxation dispersion (RD) experiments to investigate the conformational fluctuations of the RNA with atomic resolution.
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):4876-4880. doi:10.1073/pnas.0609956104
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):226-233. doi:10.1038/nsmb1059
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):1076-1083. doi:10.1038/nsmb.1494
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):786-799. doi:10.1016/j.jmb.2011.04.039
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):926-938. doi:10.1016/j.jmb.2010.10.056
Keywords: