The crystal structure of elongation factor G complexed with GDP, at 2.7 A resolution.

Abstract

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.

Functional Interactions between the G′ Subdomain of Bacterial Translation Factor EF-G and Ribosomal Protein L7/L12

“…On EF-Tu (and possibly also EF-G), this reaction is catalyzed by a conserved histidine residue (3), whose side chain is believed to rotate to a position next to the water molecule (4). In contrast to EF-Tu, EF-G contains a GЈ subdomain, which is invariably inserted between ␣-helices D G and E G (5). The GЈ subdomain is also present in the same location in EF-2, the eukaryotic cytoplasmic homolog of EF-G (6).…”

C-terminal interaction of translational release factors eRF1 and eRF3 of fission yeast: G-domain uncoupled binding and the role of conserved amino acids

“…In eukaryotes, one translational release factor, eRF1, recognizes three stop codons (class-I), and another factor, eRF3, stimulates eRF1 activity and binds guanine nucleotides (class-II)+ The mechanism by which the eRF1 protein reads the stop codon and the G protein, eRF3, controls the mode of termination have been coding and translational problems for the three decades since the discovery of the genetic code (for a review, see Nakamura et al+, 1996)+ Prokaryotes have two class-I release factors, RF1 and RF2, that recognize UAG/UAA and UGA/UAA, respectively+ From a sequence comparison of release factors of different organisms, we have proposed a model in which the class-I release factors mimic the shape of tRNA for binding to the decoding site (A site) of the ribosome and mimic a tRNA anticodon for reading the stop codon ("RF-tRNA mimicry" hypothesis; Ito et al+, 1996)+ The mimicry of tRNA by protein has been identified by means of structural studies of bacterial elongation factors EF-G and EF-Tu complexed with guanine nucleotide(s) and aminoacyl-tRNA+ The three-dimensional structure of Thermus thermophilus EF-G comprises five subdomains; the C-terminal part, domains III-V (AEvarsson et al+, 1994;Czworkowski et al+, 1994), appears to mimic the shapes of the acceptor stem, the anticodon helix, and the T stem of tRNA, respectively (Nissen et al+, 1995)+ Class-I release factors share homology with domain IV of EF-G (Ito et al+, 1996)+ Mutational studies have provided evidence that bacterial RF1 and RF2 encode a putative protein anticodon moiety (Ito et al+, 1998b; for a review, see Nakamura & Ito, 1998)+ The model of RF-tRNA mimicry predicts that a class-II factor, eRF3, may be an EF-Tu-like vehicle protein to bring class-I proteins to the A site of the ribosome+ Several lines of evidence support this view; eRF3 shows considerable C-terminal homology to EF-1a (for a review, see Stansfield & Tuite, 1994), and eRF3 and eRF1 bind in vivo and in vitro and exist as a heterodimer complex in yeast cell lysates (Stansfield et al+, 1995;Zhouravleva et al+, 1995;Ito et al+, 1998a)+ To extend the analysis of eukaryotic release factor function and interaction, we have cloned the eRF1 (identical to Sup45) and eRF3 (identical to Sup35) genes of the fission yeast Schizosaccharomyces pombe (Ito et al+, 1996(Ito et al+, , 1998a)+ These genes are homologous to Saccharomyces cerevisiae counterparts and complement temperature-sensitive mutations in sup45 and sup35 of S. cerevisiae, respectively+ S. pombe eRF3 is a protein with a molecular mass of 72+5 kDa (composed of 662 amino acids), and the deduced protein sequence of the C-terminal 430 amino acids is highly similar to that of S. cerevisiae eRF3 as well as to EF-1a+ However, the 230 N-terminal amino acids do not share any sequence homology with S. cerevisiae eRF3+ The C-terminal two-thirds are essential for viability and translation termination, while the N-terminal one-third is not conserved and is not essential ...…”

Accommodating the bacterial decoding release factor as an alien protein among the RNAs at the active site of the ribosome

“…The concept of structural molecular mimicry among protein synthesis factors and their complexes arose as a result of the X-ray structures of several factors with ligands attached. The initial example showed strikingly that the elongation factor (EF-G in bacteria) that translocates the tRNA through the ribosome was a structural mimic of the elongation factor that delivers the aminoacyl tRNA to the ribosome (EF-Tu in bacteria) when that factor was complexed with its tRNA [22][23][24][25]. The tRNA bound to EF-Tu was mimicked by domains III, IV and V of the protein structure in EF-G. A similar structural mimicry was subsequently proposed for the two classes of RFs [12], decoding RF (RF1 and RF2 in bacteria) and recycling RF (RF3 in bacteria), although at that time no structures were available.…”