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A potential RNA drug target in the hepatitis C virus internal ribosomal entry site

Published online by Cambridge University Press:  08 December 2000

ROSCOE KLINCK
Affiliation:
RiboTargets Ltd. Granta Park, Abington, Cambridge, CB1 6GB, United Kingdom
ERIC WESTHOF
Affiliation:
Institut de Biologie Moléculaire et Cellulaire/Centre National de la Recherche Scientifique, Strasbourg, F-67084, France
STEPHEN WALKER
Affiliation:
RiboTargets Ltd. Granta Park, Abington, Cambridge, CB1 6GB, United Kingdom
MOHAMMAD AFSHAR
Affiliation:
RiboTargets Ltd. Granta Park, Abington, Cambridge, CB1 6GB, United Kingdom
ADAM COLLIER
Affiliation:
RiboTargets Ltd. Granta Park, Abington, Cambridge, CB1 6GB, United Kingdom
FAREED ABOUL-ELA
Affiliation:
RiboTargets Ltd. Granta Park, Abington, Cambridge, CB1 6GB, United Kingdom
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Abstract

Subdomain IIId from the hepatitis C virus (HCV) internal ribosome entry site (IRES) has been shown to be essential for cap-independent translation. We have conducted a structural study of a 27-nt fragment, identical in sequence to IIId, to explore the structural features of this subdomain. The proposed secondary structure of IIId is comprised of two 3 bp helical regions separated by an internal loop and closed at one end by a 6-nt terminal loop. NMR and molecular modeling were used interactively to formulate a validated model of the three-dimensional structure of IIId. We found that this fragment contains several noncanonical structural motifs and non-Watson–Crick base pairs, some of which are common to other RNAs. In particular, a motif characteristic of the rRNA α-sarcin/ricin loop was located in the internal loop. The terminal loop, 5′-UUGGGU, was found to fold to form a trinucleotide loop closed by a trans-wobble U˚G base pair. The sixth nucleotide was bulged out to allow stacking of this U˚G pair on the adjacent helical region. In vivo mutational analysis in the context of the full IRES confirmed the importance of each structural motif within IIId for IRES function. These findings may provide clues as to host cellular proteins that play a role in IRES-directed translation and, in particular, the mechanism through which host ribosomes are sequestered for viral function.

Type
Research Article
Copyright
2000 RNA Society

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