The stability against chemical denaturants of the elongation factor EF-1alpha (SsEF-1alpha), a protein isolated from the hyperthermophilic archaeon Sulfolobus solfataricus has been characterized in detail. Indeed, the atypical shape of the protein structure and the unusual living conditions of the host organism prompted us to analyze the effect of urea and guanidine hydrochloride (GuHCl) on the GDP complex of the enzyme (SsEF-1alpha x GDP) by fluorescence and circular dichroism. These studies were also extended to the nucleotide-free form of the protein (nfSsEF-1alpha). Interestingly, the experiments show that the denaturation curves of both SsEF-1alpha forms present a single inflection point, which is indicative of a cooperative unfolding process with no intermediate species. Moreover, the chemically induced unfolding process of both SsEF-1alpha x GDP and nfSsEF-1alpha is fully reversible. Both SsEF-1alpha forms exhibit remarkable stability against urea, but they do not display a strong resistance to the denaturing action of GuHCl. These findings suggest that electrostatic interactions significantly contribute to SsEF-1alpha stability. Study holds ProTherm entries: 19757, 19758, 19759, 19760, 19761, 19762, 19763, 19764, 19765, 19766, 19767, 19768, 19769, 19770, 19771, 19772, 19773, 19774, 19775, 19776 Extra Details: hyperthermophilic archaeon; nucleotide-free form; cooperative unfolding; electrostatic interactions
Submitter: Connie Wang
Submission Date: April 24, 2018, 8:52 p.m.
|Number of data points||20|
|Proteins||Elongation factor 1-alpha ; Elongation factor 1-alpha|
|Assays/Quantities/Protocols||Experimental Assay: Cm ; Experimental Assay: Cm ; Experimental Assay: Cm ; Experimental Assay: Cm|
|Libraries||Mutations for sequence MSQKPHLNLIVIGHVDHGKSTLVGRLLMDRGFIDEKTVKEAEEAAKKLGKESEKFAFLLDRLKEERERGVTINLTFMRFETKKYFFTIIDAPGHRDFVKNMITGASQADAAILVVSAKKGEYEAGMSVEGQTREHIILAKTMGLDQLIVAVNKMDLTEPPYDEKRYKEIVDQVSKFMRSYGFNTNKVRFVPVVAPSGDNITHKSENMKWYNGPTLEEYLDQLELPPKPVDKPLRIPIQDVYSISGVGTVPVGRVESGVLKVGDKIVFMPAGKVGEVRSIETHHTKMDKAEPGDNIGFNVRGVEKKDIKRGDVVGHPNNPPTVADEFTARIIVVWHPTALANGYTPVLHVHTASVACRVSELVSKLDPRTGQEAEKNPQFLKQGDVAIVKFKPIKPLCVEKYNEFPPLGRFAMRDMGKTVGVGIIVDVKPAKVEIK|