Structural organization of the human eukaryotic initiation factor 5A precursor and its site-directed variant Lys50 -> Arg

The molecular properties of the human eukaryotic initiation factor 5A precu rsor and its site directed Lys50 --> Arg variant have been investigated and compared. Structure perturbation methods were used to gain information abo ut the protein architecture in solution. Intrinsic and extrinsic spectrosco pic probes strategically located in the protein matrix detected the indepen dent unfolding of two molecular regions. Three cysteines out of four were t itrated in the native protein and the peculiar presence of a tyrosinate ban d at neutral pH was detected. At alkaline pH only two tyrosines out of thre e were titratable in the native protein, with an apparent pK of about 9.9. Native protein and its Lys50 --> Arg variant reacted in a similar fashion t o guanidine and to pH variation, but differently to thermal stress. The com plex thermal unfolding of both proteins indicated the presence of intermedi ates. Spectroscopic data showed that these intermediates are differently st ructured. Consequently, the two proteins seem to have different unfolding p athways.