Effects of Thioamide Substitutions on the Conformation and Stability of α- and 310-Helices

Thiopeptides, formed by replacing the amide oxygen atom with a sp2 sulfur atom, are useful in protein engineering and drug design because they confer resistance to enzymatic degradation and are predicted to be more rigid. This report describes our free molecular dynamics simulations with explicit water and free energy calculations on the effects of thio substitutions on the conformation of α-helices, 310-helices, and their relative stability. The most prominent structural effect of thio substitution is the increase in the hydrogen bond distance from 2.1 Å for normal peptides to 2.7 Å for thiopeptides. To accommodate for the longer CS···H−N hydrogen bond, the (φ, ψ) dihedral angles of the α-helix changed from (−66°, −42°) to (−68°, −38°), and the rise per turn increased from 5.5 to 6.3 Å. For 310-helices, the (φ, ψ) dihedral angles (−60°, −20°) and rise per turn (6.0 Å) changed to (−66°, −12°) and 6.8 Å, respectively. In terms of relative stability, the most prominent change upon thio substitution is the decrease in the free energy difference, ΔA(α → 310), from 14 to 3.5 kcal/mol. Therefore, normal peptides are less likely to form 310-helix than are thiopeptides. Component analysis of the ΔA(α → 310) reviews that the entropy advantage of the 310-helix for both Ac−Ala10−NHMe and Act−Alat10−NHMe is attributed to the 310-helix being more flexible than the α-helix. Interestingly, upon thio substitution, this differential flexibility is even more apparent because the α-helix conformation of Act−Alat10−NHMe becomes more rigid due to the bulkier sulfur atom.