CONCLUSION In this study, we investigate the structural hierarchy and energy landscape of an RNA T-loop hairpin. We found that the U-turn component of the T-loop motif is ﬂexible both when the T-loop is in isolation and as part of a hairpin. The other component of the loop, the U:A trans W/H bp, can be stabilized in the context of the UA-handle motif when a stem is present. We speculate that the UA-handle plays an important role in stabilizing small RNA motifs such as the T-loop motif. Future experi- mental research in our laboratory will aim at further probing this idea. Our simulations indicate that the energy landscape for the folding of the T-loop hairpin is rugged. This roughness is likely a common feature of small RNA motifs and implies that additional help from the environment (via, for instance, interaction with neighbor- ing residues or through long range interactions) is needed to stabilize a particular conformer over another and ensure correct folding. SUPPLEMENTARY DATA Supplementary Data are available at NAR Online. ACKNOWLEDGEMENTS Andrew Jewett is thanked for helpful discussion and for the WHAM program used in data analysis. Simulations were performed using the computational resources of the California NanoSystems Institute (CNSI). L.J. wishes to dedicate this paper to St. Elizabeth Ann Seton, a great mother and teacher. David and Lucile Packard Foundation to J.-E.S., National Science Foundation (CHE-0321368 to J.-E.S., MCB-0642086 to J.-E.S. DMR-05-20415 to J.-E.S. and L.J.); National Institutes of Health (R01 GM079604-01 to L.J.). Funding to pay the Open Access publication charges for this article was provided by the National Science Foundation (NSF). Conﬂict of interest statement. None declared. REFERENCES 1. Mathews,D.H. (2006) Revolutions in RNA secondary structure prediction. J. Mol. Biol., 359, 526–532. 2. Brion,P. and Westhof,E. (1997) Hierarchy and dynamics of RNA folding. Annu. Rev. Biophys. Biomol. 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