Structural analysis of peptides encompassing all alpha-helices of three alpha/beta parallel proteins: Che-Y, flavodoxin and P21-ras: implications for alpha-helix stability and the folding of alpha/beta parallel proteins.

In an attempt to delineate the early folding events of structurally related proteins with no sequence homology, peptides including all five alpha-helices of three alpha/beta parallel open-sheet proteins, Che-Y, flavodoxin and P21-ras, have been analyzed by circular dichroism (far-UV CD) and nuclear magnetic resonance (NMR) in water and 30% (v/v) trifluoroethanol (TFE). Comparison between the helical content estimations from far-UV CD and the results from the NMR analysis renders a reasonably good qualitative correlation, indicating that the same phenomenon is underlined by both methods. Helix limits, as indicated by the existence of (i,i + 3) nuclear Overhauser effect (NOE) cross-correlations and significant up-field conformational shifts of the C alpha H protons, are practically coincident with those in the folded protein. On the other hand, the conformation of the side-chains differs markedly from those in the folded protein. Observation of NOE cross-correlations between pairs of residues at positions i,i + 3 has been used to statistically quantify free energies of i,i + 3 side-chain-side-chain interactions between the different pairs of residues in an alpha-helix. This analysis indicates that interactions between hydrophobic side-chains seem to be quite favorable for helix formation. The behaviour in aqueous solution of the structural equivalent peptides for the three proteins is quite unrelated except for the peptides corresponding to helices two and five. We postulate that, in the alpha/beta parallel proteins, those helices that join two beta-strands flanking another non-consecutive beta-strand should not be stable for folding reasons.