The Interaction between the Ribosomal Stalk Proteins and Translation Initiation Factor 5B Promotes Translation Initiation

Abstract

Ribosomal stalk proteins recruit translation elongation GTPases to the factor-binding center of the ribosome. Initiation factor 5B (eIF5B in eukaryotes and aIF5B in archaea) is a universally conserved GTPase that promotes the joining of the large and small ribosomal subunits during translation initiation. Here we show that aIF5B binds to the C-terminal tail of the stalk protein. In the cocrystal structure, the interaction occurs between the hydrophobic amino acids of the stalk C-terminal tail and a small hydrophobic pocket on the surface of the GTP-binding domain (domain I) of aIF5B. A substitution mutation altering the hydrophobic pocket of yeast eIF5B resulted in a marked reduction in ribosome-dependent eIF5B GTPase activity <i>in vitro</i> In yeast cells, the eIF5B mutation affected growth and impaired <i>GCN4</i> expression during amino acid starvation via a defect in start site selection for the first upstream open reading frame in <i>GCN4</i> mRNA, as observed with the eIF5B deletion mutant. The deletion of two of the four stalk proteins diminished polyribosome levels (indicating defective translation initiation) and starvation-induced <i>GCN4</i> expression, both of which were suppressible by eIF5B overexpression. Thus, the mutual interaction between a/eIF5B and the ribosomal stalk plays an important role in subunit joining during translation initiation <i>in vivo</i>.