Conformational Change of Elongation Factor Tu (EF-Tu) Induced by Antibiotic Binding

Abstract

Aurodox is a member of the family of kirromycin antibiotics, which inhibit protein biosynthesis by binding to elongation factor Tu (EF-Tu). We have determined the crystal structure of the 1:1:1 complex of Thermus thermophilus EF-Tu with GDP and aurodox to 2.0-Å resolution. During its catalytic cycle, EF-Tu adopts two strikingly different conformations depending on the nucleotide bound: the GDP form and the GTP form. In the present structure, a GTP complex-like conformation of EF-Tu is observed, although GDP is bound to the nucleotide-binding site. This is consistent with previous proposals that aurodox fixes EF-Tu on the ribosome by locking it in its GTP form. Binding of EF-Tu⅐GDP to aminoacyl-tRNA and mutually exclusive binding of kirromycin and elongation factor Ts to EF-Tu can be explained on the basis of the structure. For many previously observed mutations that provide resistance to kirromycin, it can now be understood how they prevent interaction with the antibiotic. An unexpected feature of the structure is the reorientation of the His-85 side chain toward the nucleotide-binding site. We propose that this residue stabilizes the transition state of GTP hydrolysis, explaining the acceleration of the reaction by kirromycin-type antibiotics.

Ribosome-induced changes in elongation factor Tu conformation control GTP hydrolysis

“…The aurodox-bound structure, shown in Fig. 2D, displays an open gate, where Ile-60 and the remainder of switch I are disordered and His-84 is repositioned toward the nucleotide (12), hinting that the increased GTPase activity induced by aurodox is achieved by gate opening. One wing of the hydrophobic gate is formed by residue Val-20, which forms part of the P loop, a conserved element in GTPases important in guanine nucleotide release (13,14).…”

Structural Basis for the Binding of Didemnins to Human Elongation Factor eEF1A and Rationale for the Potent Antitumor Activity of These Marine Natural Products

“…Competition for binding to this site accounts for the effects of the antibiotic on translation inhibition. In contrast, aurodox locks EF1A in its GTP form, even if GDP is bound to the nucleotide-binding site, 15 and prevents binding of EF1B, a GEF formerly known as EF-Ts. However, kirromycin fails to induce the GTP-like conformation of eEF1A‚GDP, which explains its inability to inhibit peptide bond formation in the eukaryotic system.…”

“…9 This rather unusual mechanism would then be similar to that of the antibiotic kirromycin, which is known to bind to prokaryotic EF1A (previously termed EF-Tu), 15 even though DB, unlike this antibiotic, does not prevent peptide bond formation. Subsequent experimental results led to the suggestion that eEF2 binding could be prevented by inhibiting eEF1A release from the ribosomal A-site, and two alternative scenarios were then envisaged: 14 (i) direct competition of the DB‚eEF1A complex with eEF2 for the same binding site on the 27) showing the role of eEF1A and eEF1B (red arrows) in translation elongation.…”

Molecular signatures in protein sequences that are characteristics of the phylum Aquificae

“…In view of the evolutionarily conserved nature of these genetic changes, it is reasonable to assume that they are modifying the cellular functions of these proteins (either their biochemical activity or their regulation) in some unique manner which is distinctive for this group of bacteria. It is of interest to note that the indel in PolA falls in a region which affects the accuracy of replication (Minnick et al, 1999), whereas that in the EF-Tu protein lies close to a region that acts as a sensor for the presence or absence of GTP and transmits this signal to other domains of the protein (Vogeley et al, 2001). The large insert in the SecA protein is also present in a region that is proximal to a nucleotidebinding fold essential for its function (Osborne et al, 2004).…”