Structural Fluctuation of the Polypeptide-Chain Elongation Factor Tu. A Comparison of Factors from Escherichia coli and Thermus thermophilus HB8

Abstract

The kinetics of hydrogen-deuterium exchange in the polypeptide chain elongation factor Tu (EF-Tu) from Escherichia coli and that from Thermus thermophilus HB8 has been examined in aqueous solutions at various pH and temperatures by means of infrared absorption measurements. The free EF-Tu from E. coli has a greater reaction rate at all pH values and at every temperature than that of the GTP-bound or GDP-bound EF-Tu. The free EF-Tu from T. thermophilus, on the other hand, has an almost equal reaction rate to that of EF-Tu · GDP in the temperature range 38–55 °C. For the peptide NH groups belonging to a medium-labile kinetic class, a small but definite difference in the rate of exchange reaction was observed between EF-Tu · GDP and EF-Tu · GTP for both E. coli and T. thermophilus. For less labile peptide NH groups, on the other hand, the rate of the exchange reaction with EF-Tu · GDP from T. thermophilus is only slightly affected by the pH of the solution at 38°C and 45°C, while the rate constant (k) with E. coli EF-Tu · GDP is pH-dependent (log k· pH). For T. thermophilus EF-Tu, heat stability measurements, kinetics of the rates of GDP and GTP dissociation, and circular dichroic measurements have also been made. The molecular basis for the thermostability of T. thermophilus EF-Tu is discussed.