Structure of HCV IRES domain II determined by NMR

Abstract

Complex RNA structures regulate many biological processes, but are often too large for structure determination by NMR methods. The 5′ untranslated region (5′ UTR) of the hepatitis C viral (HCV) RNA genome contains an internal ribosome entry site (IRES) that binds to 40S ribosomal subunits with high affinity and specificity to control translation. Domain II of the HCV IRES forms a 25-kDa folded subdomain that may alter ribosome conformation. We report here the structure of domain II as determined using an NMR approach that combines short- and long-range structural data. Domain II adopts a distorted L-shape structure, and its overall shape in the free form is markedly similar to its 40S subunit–bound form; this suggests how domain II may modulate 40S subunit conformation. The results show how NMR can be used for structural analysis of large biological RNAs.

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Figure 1: NMR structure determination of the HCV IRES domain II structure.
Figure 2: Structure determination of domain II using RDCs.
Figure 3: Solution structure of HCV IRES RNA domain II and magnesium binding sites.
Figure 4: Comparison of subdomains IIb and IIId of the HCV IRES and base pairing schemes.

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Acknowledgements

We thank E. Lau for labeled nucleotide preparation and P. Sarnow, Y. Shibata-Lukavsky, R.L. Gonzalez, D. Daniels, C. Liu and E.V. Puglisi for helpful discussions. This work was supported by grants from the US National Institutes of Health, the Hutchison Foundation and Eli Lilly. Stanford Magnetic Resonance Laboratory is supported by the Stanford School of Medicine.

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Correspondence to Joseph D Puglisi.

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Lukavsky, P., Kim, I., Otto, G. et al. Structure of HCV IRES domain II determined by NMR. Nat Struct Mol Biol 10, 1033–1038 (2003) doi:10.1038/nsb1004

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