Truncated versions of the gene for elongation factor Tu (EF-Tu; 400 aa) from the hyperthermophilic bacterium have been produced by progressive 3′→5′ trimming. The truncated genes have been expressed in and the thermal stability of the gene products has been assayed by monitoring their GDP-binding capacity after preheating the cell-free extracts at various temperatures (65-95 C). One of the truncated proteins, corresponding to the nucleotide-binding domain (G domain aa 1-200) appears to be only slightly less stable than the full-length EF-Tu. Replacement of the first 90 N-terminal residues of both the full-length EF-Tu and the isolated G domain with the corresponding sequence of the mesophilic bacterium drastically destabilizes both the complete and the truncated protein, indicating that sequence element(s) that are crucial for the attainment of a thermally stable conformation of the EF-Tu lie well within the initial portion of the G domain between residues 1 and 90. The relevant residues defy identification, however, as no amino acid preferences, or exclusive sequence element(s), appear to distinguish the N-terminal region of the thermophilic proteins from those of mesophilic counterparts. It is suggested that the thermal stability of EF-Tu is critically dependent upon unique tertiary structural interactions involving certain N-terminal residues of the molecule.


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