Bacterial translation initiation factor 2 (IF2) is a GTPase that promotes the binding of the initiator fMet-tRNA fMet to the 30S ribosomal subunit. It is often assumed that IF2 delivers fMet-tRNA fMet to the ribosome in a ternary complex, IF2GTPfMet-tRNA fMet. By using rapid kinetic techniques, we show here that binding of IF2GTP to the 30S ribosomal subunit precedes and is independent of fMet-tRNA fMet binding. The ternary complex formed in solution by IF2GTP and fMet-tRNA is unstable and dissociates before IF2GTP and, subsequently, fMet-tRNA fMet bind to the 30S subunit. Ribosome-bound IF2 might accelerate the recruitment of fMet-tRNA fMet to the 30S initiation complex by providing anchoring interactions or inducing a favourable ribosome conformation. The mechanism of action of IF2 seems to be different from that of tRNA carriers such as EF-Tu, SelB and eukaryotic initiation factor 2 (eIF2), instead resembling that of eIF5B, the eukaryotic subunit association factor. © 2010 European molecular biology organization.