Mp. Laurino et al., The beta subunit of eukaryotic translation initiation factor 2 binds mRNA through the lysine repeats and a region comprising the C-2-C-2 motif, MOL CELL B, 19(1), 1999, pp. 173-181
Eukaryotic translation initiation factor 2 (eIF2) has been implicated in th
e selection of the AUG codon as the start site for eukaryotic translation i
nitiation, since mutations in its three subunits in yeast that allow the re
cognition of a UUG codon by the anticodon of the initiator Met-tRNA(Met) ha
ve been identified. All such mutations in the beta subunit of eIF2 (eIF2 be
ta) mapped to a region containing a putative zinc finger structure of the C
-2-C-2 type, indicating that these sequences could be involved in RNA recog
nition. Another feature of eIF2 beta that could mediate an interaction with
RNA is located in the amino-terminal sequences and is composed of three re
peats of seven lysine residues which are highly conserved in other species.
We show here the ability of eIF2 beta, purified from Escherichia coli as a
fusion to glutathione S-transferase, to bind mRNA in vitro. Through a dele
tion analysis, mRNA binding was found to be dependent on the lysine repeats
and a region encompassing the C-2-C-2 motif. Strong mRNA binding in vitro
could be maintained by the presence of only one lysine or one arginine run
but not one alanine run. We further show that only one run of lysine residu
es is sufficient for the in vivo function of eIF2 beta, probably through ch
arge interaction, since its replacement by arginines did not impair cell vi
ability, whereas substitution for alanines resulted in inviable cells, mRNA
binding, but not GTP-dependent initiator Met-tRNA(Met) binding, by the eIF
2 complex was determined to be dependent on the presence of the lysine runs
of the beta subunit.