Properties of the elongation factor 1α in the thermoacidophilic archaebacterium Sulfolobus solfataricus

The elongation factor 1α (aEF‐1α) was purified to homogeneity from the thermoacidophilic archaebacterium Sulfolobus solfataricus by chromatographic procedures utilising DEAE‐Sepharose, hydroxyapatite and FPLC on Mono S. The purified protein binds [ 3 H]GDP at a 1:1 molar ratio and it is essential for poly(Phe) synthesis in vitro ; it also binds GTP but not ATP. These findings indicate that aEF‐1α is the counterpart of the eubacterial elongation factor Tu (EF‐Tu). Purified aEF‐1α is a monomeric protein with a relative molecular mass of 49 000 as determined by SDS/PAGE and by gel filtration on Sephadex G‐100; its isoelectric point is 9.1. The overall amino acid composition did not reveal significant differences when compared with the amino acid composition of eubacterial EF‐Tu from either Escherichia coli or Thermus thermophilus , of eukaryotic EF‐1α from Artemia salina or of archaebacterial EF‐1α from Methanococcus vannielii . The close similarities between the average hydrophobicity and the numbers of hydrogen‐bond‐forming or non‐helix‐forming residues suggest that common structural features exist among the factors compared. aEF‐1α shows remarkable thermophilic properties, as demonstrated by the rate of [ 3 H]GDP binding which increases with temperature, reaching a maximum at 95°C; it is also quite heat‐resistant, since after a 6‐h exposure at 60°C and 87°C the residual [ 3 H]GDP‐binding ability was still 90% and 54% of the control, respectively. The affinity of aEF‐1α for GDP and GTP was also evaluated. At 80°C K a ′ for GDP was about 30‐fold higher than K a ′ for GTP; at the same temperature K d ′ for GDP was 1.7 μM and K d ′ for GTP was 50 μM; these values were 300‐fold and 100‐fold higher, respectively, than those reported for E. coli EF‐Tu at 30°C; compared to the values at 0°C of EF‐Tu from E. coli and T. thermophilus or EF‐1α from A. salina , pig liver and calf brain, smaller differences were observed with eukaryotic factors. At 80°C k ′ −1 GDP and k ′ +1 GDP of aEF‐1α were 7.8 min −1 and 4.6 min −1 ·μM −1 , respectively; comparison with the values attributed to E. coli EF‐Tu at 30°C indicates that the dissociation rate constant increases and the association rate constant decreases, thus indicating that the higher K d ′ for GDP of aEF‐1α is due to a synchronous alteration of the rates of both the dissociation and the association processes between GDP and aEF‐1α. Besides aEF‐1α, the other two elongation factors, aEF‐2 and aEF‐1β were also present in S. solfataricus , as suggested by the occurrence of reactions typical of each of them. This allows the hypothesis that in this extremophilic archaebacterium the protein synthesis process follows the same general pattern found in all other living organisms.