Over-coordinated H-bonding in membrane proteins

A single helix from a globular protein will unfold when dissolved in an aqueous solution. But in the absence of water, the canonical H-bonds between the backbone NH at position i and backbone CO at position i−4 are the prominent stabilizing forces in α-helices. Membrane proteins, which reside in hydrophobic environments, adopt extremely stable secondary structures, which only unfold when the membrane fidelity collapses.

These canonical H-bonds responsible for stabilizing protein secondary structure are composed of a single donor and a single acceptor. Yet often, H-bond configurations involve multiple donors or acceptors. Our research investigates how polar side-chains in membrane proteins form H-bonds with over-coordinated backbone carbonyl groups which are already involved in canonical H-bonds. These types of nonconventional H-bonds are very common; 92% of all transmembrane helices have at least one of these over-coordinated H-bonds.

When polar serines exist in transmembrane helices at position i, they can self-supply their H-bonding requirements by backbonding to a backbone carbonyl group:
• 41% adopt a −60 rotamer to backbond to the i−4 carbonyl.
• 26% adopt a +60 rotamer to backbond to the i−3 carbonyl.
• 31% adopt a ±180 rotamer to backbond to their own carbonyl.
Interestingly, these serine side-chains are sometimes over-coordinated to both the i−3 and the i−4 backbone carbonyls simultaneously, creating a multiplexed H-bonding system with multiple proton donors and acceptors.

Using fourier transform infrared spectroscopy combined with density functional theory quantum simulations, we measure these over-coordinated H-bond strengths. We find that they can be stronger than canonical H-bonds by over 50%.