Question

NMR measurements have shown that poly-L-lysine is a random coil at pH=7 but becomes alpha helical as the pH is raised above 10.

a) Why can NMR give this sort of information? (what exactly does NMR look at?)

b) what may be causing this pH-dependent transition?

c) predict the pH dependence of a helix-coil transition if the poly-L-aspartate

Answer 1

(a) Electrostatic repulsion between positively charged ϵ-amino groups hinders α-helix formation at pH 7. At pH 10, the side chains become deprotonated, allowing α-helix formation.

The alpha helix (α-helix) is a commonsecondary structure of proteins and is a righthand-coiled or spiral conformation (helix) in which every backbone N-Hgroup donates a hydrogen bond to the backbone C=O group of the amino acidfour residues earlier (hydrogen bonding). This secondary structure is also sometimes called a classic Pauling–Corey–Branson alpha helix (see below). The name3.6₁₃-helix is also used for this type of helix, denoting the number of residues per helical turn, and 13 atoms being involved in the ring formed by the hydrogen bond. Among types of local structure in proteins, the α-helix is the most regular and the most predictable from sequence, as well as the most prevalent.

(b) Poly-l-glutamate is a random coil at pH 7 and becomes α helical below pH 4.5 because the γ-carboxylate groups become protonated.

C) poly L-glutamic acid becomes alpha helical at very low pH

Protein folding and conformational changes are influenced by protein-water interactions and, as such, the energetics of protein function are necessarily linked to water activity. Here, we have chosen the helix-coil transition in poly(glutamic acid) as a model system to investigate the importance of hydration to protein structure by using the osmotic stress method combined with circular dichroism spectroscopy. Osmotic stress is applied using poly(ethylene glycol), molecular weight of 400, as the osmolyte. The energetics of the helix-coil transition under applied osmotic stress allows us to calculate the change in the number of preferentially included water molecules per residue accompanying the thermally induced conformational change. We find that osmotic stress raises the helix-coil transition temperature by favoring the more compact α-helical state over the more hydrated coil state. The contribution of other forces toα-helix stability also are explored by varying pH and studying a random copolymer, poly(glutamic acid-r-alanine)