Thermodynamics of GTP and GDP Binding to Bacterial Initiation Factor 2 Suggests Two Types of Structural Transitions

Thermodynamics of the GTP-GDP-operated Conformational Switch of Selenocysteine-specific Translation Factor SelB

“…, which is larger than that estimated for EF-G (35) and IF2 (34), 250 and 290 cal ϫ mol Ϫ1 ϫ K Ϫ1 , respectively. Part of this rearrangement was suggested to come from alterations in the switch 1 and switch 2 regions of the factors, by analogy to the well documented structural changes observed in EF-Tu and other GTPases (36).…”

“…In fact, crystal structures suggested that the overall domain arrangements in SelB (13), IF2/eIF5B (30), or EF-G (31-33) are essentially identical in the apo form and GDP-and GDPNP-bound forms; the observed limited conformational changes in the switch 1 and 2 regions were confined to the immediate vicinity of the ␥-phosphate. In contrast, the ITC results indicate a large difference between the GTP-bound and apo forms, corresponding to rearrangements upon GTP binding of about 42 amino acids in SelB, 58 amino acids in IF2 (34), and 19 amino acids in EF-G (35). The use of GDPNP, rather than GTP, in the structural work might explain these discrepancies.…”

Thermodynamic Characterization of ppGpp Binding to EF-G or IF2 and of Initiator tRNA Binding to Free IF2 in the Presence of GDP, GTP, or ppGpp

“…The affinities of IF2 for ppGpp (Table 1) and GDP were similar and about 2.5-fold to 5-fold higher than the affinity of IF2 for GTP. 15 The affinities of EF-G for ppGpp (Table 1, Fig. 2b), GDP, and GTP were similar, especially at physiologically relevant temperatures (i.e., 25-37°C) ( Table S1 and Hauryliuk et al 14 ).…”

“…K a is the affinity constant (SD did not exceed ±20%); K d is the dissociation constant (calculated as 1/K a ); ΔH is the enthalpy variation (SD did not exceed ±10%); TΔS is the entropy variation (calculated from the equation ΔG = ΔH − TΔS); and ΔG is the Gibbs energy (calculated from the equation ΔG = − RTlnK a ). GTP to IF2 are − 577 ± 23 and − 858 ± 25 cal mol − 1 K − 1 , respectively 15 (Table 2), and the corresponding estimates for the binding of GDP or GTP to EF-G are − 21 ± 5 or − 270 ± 20 cal mol − 1 K − 1 , respectively 14 ( Table 2). On the assumption of the additive contributions of the different phosphor groups to the ΔC p values, 21 we found the contribution of γ-phosphate on GTP to be similar in IF2 (− 281 ± 34 cal mol − 1 K − 1 ) and EF-G (− 249 ± 21 cal mol − 1 K − 1 ).…”

“…Recently, we used isothermal titration calorimetry (ITC) to characterize the thermodynamics of interaction between G nucleotides and the translational GTPases EF-G, 14 IF2, 15 and eukaryotic release factor 3. 16 For these interactions, we determined the equilibrium dissociation constant (K d ), which provides the Gibbs standard free energy of the interaction (ΔG°), as well as enthalpy (ΔH°) and entropy (ΔS°).…”