04 Protein structure and function

4 Protein structure and function

To understand the relevance of protein structure for protein function.
To understand the molecular interactions that determine the shape of proteins.
To appreciate the dynamic nature of proteins and their denaturation by heat and acid

Levels of protein structure
Interactions that stabilise protein structure
Active site
Induced fit

Although this lecture series focuses on nutrients and their metabolism, we will come across proteins in many forms, such as enzymes, receptors, transporters, peptide hormones etc.

A basic understanding of proteins is important to understand nutrient metabolism. Let us have a look at some proteins (Fig. 1) and watch them in action (video below).

Several important concepts of protein structure can be appreciated just by looking carefully at Fig. 1. There are simple proteins that contain only one protein chain (a single colour, encoded by one gene). Others are complex and are comprised of several chains (different colours and encoded by several genes). Ribosomes (at the bottom of Fig. 1) are even a mix of RNA and proteins. Most proteins have a globular appearance, which suggests that a compact structure is a favourable arrangement.

All proteins are made up of the 20 amino acids, which you encountered in chapter 1. Why are proteins not unwinding into a long thread of amino acids? The linear arrangement of amino acids is held together by covalent bonds, which are very strong. Even when boiled these bonds do not break. The string of amino acids making up a protein is called the primary structure.

The bond between two amino acids is called the peptide bond and is generated by elimination of water when amine of one amino acid reacts with the carboxyl-goup of another amino acid. Breakdown of peptides thus occurs by hydrolysis. Inspection of any peptide reveals atoms that are polarised. The carbonyl-oxygen (C=O) is partially negative and the hydrogen associated with the nitrogen is partially positive. Hydrogen bonds (dotted) can form between these groups: =N-H…O=C if they are in close proximity.

When the peptide is threaded like a helix and one turn takes about 4 amino acids, the carbonyl-oxygen (C=O) can form a hydrogen bond (yellow in Fig. 3) with a Nitrogen-Hydrogen (N-H) of the residue four positions further on or behind. This structure is very stable, because the amino acids do not need to find other partners to interact with. As a result, most proteins have a good proportion of amino acids forming alpha-helices. Important to note: In an alpha-helix side chains point away from the centre of the helix (Fig. 3) avoiding steric problems. Another structure that proteins can form readily is the beta-sheet (Fig. 4).

Instead of hydrogen bonding within the structure, hydrogen bonds are here formed between sheets. Alpha-helix and beta-sheet are called secondary structures. There are more secondary structures, particular the turns at the end of helices or sheets. The tertiary structure of a protein is the complete folding of a primary structure into the final protein structure. Often protein folding is similar in different, sometimes unrelated proteins. Such common protein folds illustrate the multiple uses of successful scaffolds to form globular structures with a couple of key residues in the active site.

Let us look at the example of triosephosphate isomerase, one of the enzymes involved in glycolysis (Fig. 5).

The left presentation shows the surface of the protein as you would see it at very high magnification. Highlighted are the two chains of the protein forming a dimer of identical subunits. An arrangement of several peptide chains forming a functional protein is called the quarternary structure. In the center panel the secondary structures are highlighted. Helices are in blue, beta sheets in magenta and turns in red. On the right the primary structure is depicted, which is rather confusing. The yellow hue comes from highlighting hydrogen bonds. While you cannot see individual ones, it illustrates how many there are to hold the proteins together. Hydrogen bonds are depicted in more detail in Fig. 3 and 4. Hydrogen bonds are one reason that proteins do not unwind spontaneously. However, there are more interactions that hold proteins together (Fig. 6). They are called weak interactions, because the bond energy is much lower than that of a covalent bond.

You already appreciate the role of hydrogen bonds in holding secondary structures together, but they also are important to stabilise tertiary and quarternary structures. Another important interaction is the hydrophobic effect. It is based on the gain of entropy when water molecules are released from the core of the protein. Non-polar sidechains are typically found in the core of a protein, which cannot form hydrogen bonds with water molecules. If water molecules were present in the core they would have to from concatenated structures of hydrogen-bonded water molecules with little mobility, which is a state of low entropy. Although fixed water molecules are rarely found inside proteins, high resolution structures show quite a few water molecules tightly bound to the surface of proteins (Fig. 7).

In addition, charged sidechains can form ionic bonds. Hydrogen bonds can form between polar hydrogen-containing bonds of carbon, nitrogen and oxygen to other atoms with a free electron pair within a protein or between protein subunits. Cation-pi interactions form because aromatic rings are electron-rich structures that can interact with electron poor positively charged sidechains. These weak interactions can be broken by heating or addition of acid, in which case the protein denatures (see video below). Denatured proteins look white and curdled. Denaturation of protein occurs in the acid environment of the stomach and facilitates the access of proteases to hydrolyse peptide bonds.

You have now a good idea how proteins are held together. One of the main functions of the three-dimensional protein structure is to bring critical residues together that form the active site of an enzyme.

Question

Answer

Question

Answer

In Fig. 8 you can see key residues (with reactive atoms in blue and red) forming the active site of triosephosphate isomerase. They emanate from different helices and sheets of the protein. The substrate (purple) is located in the centre of the active site.

We often imagine proteins as rigid and the substrates colliding with them. When we cover enzymatic catalysis you will see that interactions between the substrate and the enzyme are critical to achieve a substrate conformation which facilitates a chemical reaction (Chapter 5). This often includes some conformational change of the protein, as well. This is particular obvious in the case of hexokinase, which encloses glucose during catalysis. This mechanism is called induced fit (Fig. 9). Induced fit is particularly important in the catalysis of membrane transport (Chapter 6).

The primary structure of a proteins is the linear chain of amino acids connected by peptide bonds
Secondary structures are typical structural elements that are readily formed by a chain of amino acids and are found in many proteins. Typical secondary structures are alpha-helix, beta-sheet and turns.
The tertiary structure is the unique shape adopted by a full-length protein chain.
The quarternary structure refers to a complex of two or more protein chains making up a larger protein complex.
Secondary, tertiary and quaternary structures are held together by different molecular interactions, such as hydrogen bonds, hydrophobic effect, ionic interactions, cation-pi interactions and disulfide bridges.
While covalent bonds cannot be broken by heat and treatment with acid, secondary, tertiary and quarternary structures are denatured, resulting in curdling of the protein. This is an important part of protein digestion in the stomach. 
The precise tertiary structure is important to align all residues that are critical for catalysis.
Ligands and protein have dynamic interactions, which changes the structure of the ligand, but also changes the structure of the protein.

Looking for a quick review of what you learned in this chapter, watch the video below.