Identification of a nucleic acid binding domain in eukaryotic initiation factor eIFiso4G from wheat.
Higher plants have two complexes that bind the m7G-cap structure of mRNA and mediate interactions between mRNA and ribosomal subunits, designated eIF4F 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 purified by affinity chromatography. We show here that eIFiso4G contains a central protease-resistant domain that binds specifically to nucleic acids. This domain spans Gln170 to Glu443 and includes four of the six homology blocks shared by eIFiso4G and eIF4G. A slightly shorter overlapping sequence, from Gly202 to Lys445, had no nucleic acid binding activity, indicating that the N-terminal end of the nucleic acid binding site lies within Gln170 to Arg201. 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 double-stranded sequences.
Kim
C Y
CY
Department of Biochemistry, University of California, Riverside, California 92521, USA.
Takahashi
K
K
Nguyen
T B
TB
Roberts
J K
JK
Webster
C
C
D000595:Amino Acid Sequence; D000818:Animals; D001665:Binding Sites; D002482:Cellulose / Q000031:analogs & derivatives / Q000378:metabolism; D002851:Chromatography, High Pressure Liquid; D016384:Consensus Sequence; D039603:Eukaryotic Initiation Factor-4G; D006801:Humans; D013058:Mass Spectrometry; D008969:Molecular Sequence Data; D009696:Nucleic Acids / Q000378:metabolism*; D009838:Oligodeoxyribonucleotides / Q000378:metabolism; D010448:Peptide Initiation Factors / Q000378:metabolism*; D012314:RNA Cap Analogs / Q000378:metabolism; D012315:RNA Caps / Q000378:metabolism; D012333:RNA, Messenger / Q000378:metabolism; D011817:Rabbits; D012685:Sepharose / Q000378:metabolism; D014908:Triticum / Q000378:metabolism*; D003313:Zea mays
D039603:Eukaryotic Initiation Factor-4G; D009696:Nucleic Acids; D009838:Oligodeoxyribonucleotides; D010448:Peptide Initiation Factors; D012314:RNA Cap Analogs; D012315:RNA Caps; D012333:RNA, Messenger; C021722:oligo(dT)-cellulose; C017516:7-methylguanosine triphosphate; D002482:Cellulose; D012685:Sepharose; C008881:phosphocellulose