Studies on the polypeptide elongation factor 2 from Sulfolobus solfataricus. Interaction with guanosine nucleotides and GTPase activity stimulated by ribosomes.
The elongation factor 2 from the thermoacidophilic archaeon Sulfolobus solfataricus (SsEF-2) binds [3H]GDP at 1:1 molar ratio. The bound [3H]GDP is displaced by GTP or its nonhydrolyzable analogue guanyl-5'-yl imidodiphosphate (Gpp(NH)p) but not by ATP, thus indicating that only the two guanosine nucleotides compete for the same binding site. The affinity of SsEF-2 for [3H]GDP is higher than that for GTP and Gpp(NH)p. On the contrary, in the presence of ribosomes the affinity of SsEF-2 for GDP is lower than that for Gpp(NH)p. SsEF-2 is endowed with an intrinsic hardly detectable GTPase activity that is stimulated by ribosomes up to 2000-fold. The ribosome-stimulated SsEF-2 GTPase (GTPaser) reaches a maximum at pH 7.8 and is not affected by ATP but is competitively inhibited by either GDP or Gpp(NH)p. Both Km for [gamma-32P]GTP and kcat of GTPaser increase with increasing temperature, and the highest catalytic efficiency is reached at 80 degrees C. The ADP-ribosylation of SsEF-2 does not significantly affect either the binding of GDP and GTP or the kinetics of the GTPaser. A hypothesis on the stimulation by ribosome of SsEF-2 GTPase is proposed.
Raimo
G
G
Dipartimento di Biochimica e Biotecnologie Mediche, Università di Napoli Federico II, Italia.
Masullo
M
M
Bocchini
V
V
D000246:Adenosine Diphosphate Ribose / Q000378:metabolism; D002384:Catalysis; D004789:Enzyme Activation; D020963:GTP Phosphohydrolase-Linked Elongation Factors / Q000378:metabolism*; D006150:Guanine Nucleotides / Q000378:metabolism*; D006358:Hot Temperature; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D020652:Peptide Elongation Factor 2; D010445:Peptide Elongation Factors / Q000378:metabolism*; D012270:Ribosomes / Q000378:metabolism*; D016944:Sulfolobus / Q000201:enzymology / Q000378:metabolism*
D006150:Guanine Nucleotides; D020652:Peptide Elongation Factor 2; D010445:Peptide Elongation Factors; D000246:Adenosine Diphosphate Ribose; D020963:GTP Phosphohydrolase-Linked Elongation Factors