PMID:2223877
| Citation |
Pieper, U, Ehbrecht, HJ, Fliess, A, Schick, B, Jurnak, F and Pingoud, A (1990) Genetic engineering, isolation and characterization of a truncated Escherichia coli elongation factor Tu comprising domains 2 and 3. Biochim. Biophys. Acta 1087:147-56 |
|---|---|
| Abstract |
A deletion mutant of a plasmid born Escherichia coli tufA gene, which codes for a truncated elongation factor Tu comprising domains 2 and 3, has been constructed by genetic engineering. This gene was overexpressed in E. coli, and a polypeptide representing the truncated elongation factor Tu was isolated, purified to near homogeneity, crystallized and characterized physico-chemically as well as biochemically. Circular dichroism spectroscopy and limited tryptic digestion demonstrate that the isolated domain pair 2 and 3 behaves like an independent folding unit which adopts a similar secondary and most likely, tertiary, structure to that present in the intact elongation factor Tu. However, the isolated domain pair 2 and 3 does not interact with aminoacyl-tRNA or the antibiotic kirromycin, two ligands which were shown previously by cross-linking experiments to be in contact with amino acid residues located in domains 1 and 2, and domain 3, respectively. The results suggest that the isolated domain pair 2 and 3 by itself forms too few contacts with these ligands to form a stable complex. Furthermore, the data suggest that domain 1 in intact EF-Tu, in a subtle but nevertheless decisive manner, alters the conformation of the other two domains in such a way that all three domains cooperatively create a high affinity binding site for aminoacyl-tRNA and the antibiotic kirromycin. |
| Links | |
| Keywords |
Circular Dichroism; Cloning, Molecular; Crystallization; Electrophoresis, Polyacrylamide Gel; Escherichia coli/genetics; Gene Expression Regulation, Bacterial; Genes, Bacterial; Guanidine; Guanidines/pharmacology; Mutation; Peptide Elongation Factor Tu/genetics; Peptide Elongation Factor Tu/isolation & purification; Peptide Elongation Factor Tu/metabolism; Protein Conformation; Protein Denaturation; Pyridones/metabolism; RNA, Transfer, Amino Acyl/metabolism; Solubility; Temperature |
| edit table |
Significance
You can help EcoliWiki by summarizing why this paper is useful
Useful Materials and Methods
You can help Ecoliwiki by describing the useful materials (strains, plasmids, antibodies, etc) described in this paper.
Annotations
<annotationlinks/>
EcoliWiki Links
Add links to pages that link here (e.g. gene, product, method pages)
References
See Help:References for how to manage references in EcoliWiki.