Identification of a nucleic acid binding domain in eukaryotic initiation factor eIFiso4G from wheat

Higher plants have two complexes that bind the m (7)G-cap structure of mRNA and mediate interactions between mRNA and ribosomal subunits, designated e IF4F and eIFiso4F. Both complexes contain a small subunit that binds the 5' -cap structure of mRNA, and a large subunit, eIF4G or eIFiso4G, that binds other translation factors and RNA. Sequence-specific proteases were used to cleave native cap-binding complexes into structural domains, which were pu rified by affinity chromatography. We show here that eIFiso4G contains a ce ntral protease-resistant domain that binds specifically to nucleic acids. T his domain spans Gln(170) to Glu(443) and includes four of the six homology blocks shared by eIFiso4G and eIF4G. A slightly shorter overlapping sequen ce, from Gly(202) to Lys(445), had no nucleic acid binding activity, indica ting that the N-terminal end of the nucleic acid binding site lies within G ln(170) to Arg(201). The binding of the central domain and native eIFiso4F to RNA homopolymers and double- and single-stranded DNAs was studied. Both molecules had highest affinity for poly(G) and recognized single- and doubl e-stranded sequences.