Elongation factor G (EF‐G) catalyzes the translocation step of protein synthesis in bacteria, and like the other bacterial elongation factor, EF‐Tu‐‐whose structure is already known‐‐it is a member of the GTPase superfamily. We have determined the crystal structure of EF‐G‐‐GDP from Thermus thermophilus. It is an elongated molecule whose large, N‐terminal domain resembles the G domain of EF‐Tu, except for a 90 residue insert, which covers a surface that is involved in nucleotide exchange in EF‐Tu and other G proteins. The tertiary structures of the second domains of EF‐G and EF‐Tu are nearly identical, but the relative placement of the first two domains in EF‐G‐‐GDP resembles that seen in EF‐Tu‐‐GTP, not EF‐Tu‐‐GDP. The remaining three domains of EF‐G look like RNA binding domains, and have no counterparts in EF‐Tu.

Analysis of nucleotide binding induced conformational changes in the current and previous HslU structures suggests a protein unfolding-coupled translocation mechanism. In this mechanism, unfolded polypeptides are threaded through the aligned pores of the ATPase and peptidase and translocated into the peptidase central chamber.

The observed nucleotide-dependent conformational changes in HslU and their governing principles provide a framework for the mechanistic understanding of other AAA(+) proteins.