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Volume 142, Issue 9

Manipulation of the gene provides clues to the localization of sequence element(s) involved in the thermal stability of elongation factor Tu Free

Abstract

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.

  • Received:
  • Accepted:
  • Revised:
  • Published Online:
Keyword(s): elongation factor Tu , extreme thermophiles , genetic manipulation , protein engineering and thermostability
* Society for General Microbiology 1996
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1996-09-01
2024-04-18
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Manipulation of the tuf gene provides clues to the localization of sequence element(s) involved in the thermal stability of Thermotoga maritima elongation factor Tu
Microbiology 142, 2525 (1996); https://doi.org/10.1099/00221287-142-9-2525
/content/journal/micro/10.1099/00221287-142-9-2525
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