The Basics of Protein Folding

Proteins fold into shapes that allow them to perform a variety of jobs. The classic principle, based on Christian Anfinsen’s experiments, is that all the information for the native three-dimensional conformation of a protein is encoded in its amino acid sequence.


However, proteins are prone to misfolding, which leads to disease states such as cystic fibrosis and Tay-Sachs disease.

Amino Acid Sequence

Proteins are large biological molecules that fold into functional three dimensional structures essential for numerous biochemical processes. The amino acid sequence in a protein determines its native structure. A protein can exist in several different states, each characterized by distinct biophysical properties.

Each state is associated with a different energy cost, and the favored protein state depends on the chemistry of the surrounding environment, such as pH, salt concentration and temperature. Alterations in the protein’s environment can cause it to misfold or unfold, a process that results in loss of function and is referred to as protein denaturation.

Once proteins have a folded three-dimensional structure, they are stabilized by a number of interactions including hydrogen bonds (the attraction between unlike electric charges between atoms in the protein’s backbone) and ionic interactions (the interaction between electrostatic opposites of charged residues). Disulfide bridges (covalently linked sulfur-sulfur chemical bonds between adjacent cysteine amino acids) also help to stabilize the protein’s tertiary structure.

A protein can have an astronomical number of different possible conformations and each one has its own energy cost. The process of protein folding occurs within a configuration space described by Joseph Bryngelson and Peter Wolynes as the protein’s energy landscape. It is believed that natural selection has optimized the protein’s energy landscape so that the favored protein state has the lowest possible Gibbs free energy value.

Hydrophobic and Polar Amino Acids

The hydrophobic interactions of amino acid side chains play a significant role in protein folding. Those amino acids with hydrophobic side chains tend to be buried in the interior of a protein to minimise contact with water. These are called non-polar amino acids. Those with polar side chains and charged R-groups, on the other hand, can interact strongly with water molecules to form hydrogen bonds. These are called hydrophilic amino acids.

Amino acids with polar side chains can also form ionic interactions with oppositely charged amino acid side chains, or with other polar amino acids (or other proteins). This is important for stabilising three-dimensional protein structures.

Moreover, polar amino acids are often found on the surface of a protein where they can form intermolecular hydrogen bonds. This is essential for the formation of alpha helix and beta sheet structures.

Finally, polar amino acids can also form intramolecular hydrogen bonds, as well as ionic interactions and disulfide bridges (covalently linked cysteine residues). This is essential for the formation of tertiary structure.

In the framework model, amino acids with polar and hydrophobic properties are arranged in a parallel manner in the core of a protein to form alpha helix and beta sheet structures. The resulting proteins are highly stable and have low time correlation functions (the probability that a protein adopts its native state) for a random sequence of amino acids. This is because the protein folds through a series of stable intermediate states.

Van der Waals Interactions

The hydrophobic amino acids tucked away in the core of proteins form weak Van der Waals interactions. These forces aren’t the strongest of the forces involved in protein folding, but they contribute to stabilizing the protein tertiary structure. The polar amino acid side chains that face the outside of the protein can also form hydrogen bonds with water molecules and with other polar amino acids. These bonds are stronger than the Van der Waals interactions.

During protein folding, buried amino acid side chains can interact with each other through electrostatic forces. These electrostatic interactions are a minor contributor to the increase in conformational entropy associated with folding, and they tend to be less favorable than hydrogen bonding.

Another important force that helps to stabilize the tertiary structure of a protein is the formation of ionic interactions between amino acid side chains that have full positive or negative charges. These interactions can form attractive salt bridges that help to stabilize the protein, but they are usually not as favorable as hydrogen bonding.

Hydrogen bonds between donor and acceptor atoms of a protein can be formed in either the native or denatured state of the protein. The hydrogen bonds in the denatured protein are more favorable than those formed in the native protein, because the atoms of the donor and acceptor proteins are more tightly packed together in the denatured protein. This tight packing of the atoms reduces the free energy of the hydrogen bond, and so the denatured protein has more entropy than the native protein.

Hydrogen Bonds

Proteins fold from an unfolded, randomly coiled state into their characteristic three dimensional structure. During this process, hydrogen bonds form between the amino acid side chains and water molecules in the hydration layer that covers the protein surface. Hydrogen bonding, along with ionic interactions, dipole-dipole interactions and London dispersion forces, contributes to the tertiary structure of proteins. During protein folding, the amino acids with nonpolar R groups cluster together on the inside of the protein leaving the polar, hydrophilic amino acids to interact with each other through hydrogen bonding and other van der Waals interactions (see this scrollable interactive from LabXchange for more).

The protein hydration layer is a key factor in the thermodynamics of the protein folding process. During the folding process, a number of enthalpic and entropic costs are incurred by the protein and the hydration solvent (see this graphic from Science Direct). The protein and hydration solvent must balance these against each other in order for the protein to reach its native state.

In general, hydrogen bonds are formed between carbonyl oxygen atoms in the protein backbone and the corresponding hydrogen atoms in water molecules that form part of the hydration layer surrounding the protein. Since the water molecule is able to establish a hydrogen bond with both the carbonyl oxygen and the hydrogen of the amine groups, the water plays a critical role in the formation and stabilization of the protein structure.