Thermodynamic properties of nucleotide-free EF-Tu from Thermus thermophilus in the presence of low-molecular weight effectors of its GTPase activity

Abstract

The thermal transition of elongation factor EF-Tu from Thermus thermophilus in the presence of low-molecular weight effectors was studied by differential scanning calorimetry. The effectors of GTPase activity used were the antibiotic kirromycin and the cations Li⁺, Na⁺, K⁺ and NH₄⁺ in the chloride form. The temperature of thermal denaturation and the cooperativity of the transition of nucleotide-free EF-Tu (EF-Tu_f) in the presence of kirromycin are comparable with those of the EF-Tu·guanosine-5′-[β,γ-imido]triphosphate (GppNHp) form, indicating similar conformational states. Increased concentrations of Na⁺ and K⁺ stabilized EF-Tu_f in a manner similar to GppNHp. NH₄⁺ decreased the transition temperature of EF-Tu_f and Li⁺ decreased both the temperature and the calorimetric enthalpy of the thermal transition of EF-Tu_f. In the presence of salts, binding of kirromycin had a stabilizing effect on EF-Tu_f. Correlation between the GTPase activity and thermodynamic characteristics of EF-Tu_f induced by kirromycin in the absence or presence of the cations is discussed.