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Molecular and Cellular Biology, August 2000, p. 6019-6029, Vol. 20, No. 16
Department of Microbiology and Immunology,
State University of New York Health Science Center at Brooklyn,
Brooklyn, New York 11203,1 and A.
N. Belozersky Institute of Physico-Chemical Biology, Moscow State
University, 119899 Moscow, Russia2
Received 14 February 2000/Returned for modification 29 March
2000/Accepted 22 May 2000
Mammalian eukaryotic initiation factor 4GI (eIF4GI) may be divided
into three similarly sized regions. The central region (amino acids
[aa] 613 to 1090) binds eIF3, eIF4A, and the encephalomyocarditis virus (EMCV) internal ribosomal entry site (IRES) and mediates initiation on this RNA. We identified the regions of eIF4GI that are
responsible for its specific interaction with the IRES and that are
required to mediate 48S complex formation on the IRES in vitro.
Mutational analysis demarcated the IRES binding fragment of eIF4GI (aa
746 to 949) and indicated that it does not resemble an RNA recognition
motif (RRM)-like domain. An additional amino-terminal sequence
(aa 722 to 746) was required for binding eIF4A and for 48S complex
formation. eIF4GI bound the EMCV IRES and
0270-7306/00/$04.00+0
Copyright © 2000, American Society for Microbiology. All rights reserved.
Physical Association of Eukaryotic Initiation Factor 4G (eIF4G)
with eIF4A Strongly Enhances Binding of eIF4G to the Internal
Ribosomal Entry Site of Encephalomyocarditis Virus and Is
Required for Internal Initiation of Translation
-globin mRNA with similar affinities, but association with eIF4A
increased its affinity for the EMCV IRES (but not
-globin RNA) by 2 orders of magnitude. On the other hand, eIF4GI mutants with defects in binding eIF4A were defective in mediating 48S complex formation even if
they bound the IRES normally. These data indicate that the eIF4G-eIF4A
complex, rather than eIF4G alone, is required for specific
high-affinity binding to the EMCV IRES and for internal ribosomal entry
on this RNA.
*
Corresponding author. Mailing address: Department of
Microbiology and Immunology, State University of New York Health
Science Center at Brooklyn, 450 Clarkson Ave., Box 44, Brooklyn, NY
11203. Phone: (718) 270-1034. Fax: (718) 270-2656. E-mail:
tpestova{at}netmail.hscbklyn.edu.
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