Proteins

1. Overview

Proteins are polymers built from amino acid monomers joined by peptide bonds. The order of amino acids (the primary structure) determines how a chain coils and folds into its secondary, tertiary and, in many proteins, quaternary structure. These higher levels are held in place by hydrophobic interactions, hydrogen bonds, ionic bonds and covalent disulfide bonds. The final three-dimensional shape decides whether a protein is a soluble globular protein with a physiological role, such as haemoglobin, or an insoluble fibrous protein with a structural role, such as collagen.

Key Definitions

• Amino acid: the monomer of proteins, containing a central carbon bonded to an amino group, a carboxyl group, a hydrogen atom and a variable R-group.
• Peptide bond: the covalent bond formed between the carboxyl group of one amino acid and the amino group of another in a condensation reaction.
• Primary structure: the specific sequence and number of amino acids in a polypeptide chain.
• Secondary structure: the regular coiling or folding of the polypeptide backbone into alpha-helices or beta-pleated sheets, held by hydrogen bonds.
• Tertiary structure: the overall three-dimensional shape of a single polypeptide, held by hydrophobic interactions, hydrogen bonds, ionic bonds and disulfide bonds.
• Quaternary structure: the structure formed when two or more polypeptide chains, sometimes with a non-protein group, associate to make a functional protein.
• Globular protein: a protein folded into a compact, roughly spherical shape that is generally soluble and has a physiological role.
• Fibrous protein: a long, narrow protein that is generally insoluble and has a structural role.
• Haem group: an iron-containing prosthetic group, one per polypeptide, that binds reversibly to one oxygen molecule in haemoglobin.

Content

Amino acid structure

Every amino acid is built around a central carbon atom (the alpha-carbon). Bonded to this carbon are four groups: an amino group ($\text{-NH}_2$), a carboxyl group ($\text{-COOH}$), a single hydrogen atom, and a variable R-group (side chain). The R-group is the only part that differs between the 20 amino acids found in proteins; it is what gives each amino acid its chemical character, for example whether it is acidic, basic, polar or hydrophobic. A useful general formula is:

$$\text{H}_2\text{N-CHR-COOH}$$

Formation and breakage of a peptide bond

A peptide bond forms between the carboxyl group of one amino acid and the amino group of the next. This is a condensation reaction: a molecule of water is removed, and the C–N bond that joins the two residues is the peptide bond. The product of two joined amino acids is a dipeptide; many joined together form a polypeptide.

The reverse process is hydrolysis: a molecule of water is added across the peptide bond to break it, releasing the individual amino acids again. This happens during digestion of dietary protein.

Primary structure

The primary structure is the precise sequence and number of amino acids in a polypeptide, read from one end to the other. It is determined by the gene that codes for the protein. The primary structure is the most important level because the position of every R-group fixes how and where the chain can fold; a change of a single amino acid can change the whole shape and therefore the function.

Secondary structure

Once the chain exists, hydrogen bonds form along the backbone between the parts of the peptide bonds themselves. Specifically, the slightly negative C=O (carbonyl) group of one peptide bond attracts the slightly positive N–H group of another peptide bond further along the chain. These regular hydrogen-bonding patterns pull the chain into two common shapes:

• the alpha-helix, a right-handed coil;
• the beta-pleated sheet, where sections of the chain lie side by side.

Note that these bonds involve the backbone of the peptide bonds, not the R-groups (those interact later, in the tertiary structure). Although each hydrogen bond is individually weak, there are very many of them, so secondary structures are stable.

Tertiary structure

The tertiary structure is the overall, unique three-dimensional shape of a single polypeptide, produced when the chain folds further so that R-groups can interact. This shape is held by four types of interaction (summarised in the table below). The tertiary structure is what gives globular proteins, including enzymes and haemoglobin chains, their specific functional shape.

Quaternary structure

A quaternary structure exists only in proteins made of two or more polypeptide chains held together by the same kinds of interaction. The structure may also include a non-protein (prosthetic) group. Haemoglobin, with four chains and four haem groups, is the standard example.

Interactions that hold proteins in shape

Four types of interaction maintain tertiary and quaternary structure, all acting between R-groups. The table below summarises where each one acts and what disrupts it:

Interaction	Acts between	Relative strength	What breaks it
Hydrophobic interactions	non-polar (water-hating) R-groups, clustered in the core away from water	weak individually	non-polar solvents; heat
Hydrogen bonding	polar R-groups	weak (but numerous)	high temperature; extremes of pH
Ionic bonding	oppositely charged R-groups (one $+$, one $-$)	moderate	changes in pH (alters the charges)
Disulfide bonds (covalent, $\text{-S-S-}$)	the sulfur atoms of two cysteine R-groups	strongest	reducing agents; not broken by mild heating

The clustering of hydrophobic R-groups in the core also pushes hydrophilic R-groups to the outside of the molecule, which is what tends to make globular proteins soluble.

Globular and fibrous proteins

Globular proteins fold into a compact, roughly spherical shape with hydrophilic R-groups on the outside and hydrophobic R-groups buried inside; this makes them generally soluble in water and suited to physiological (metabolic) roles such as transport, catalysis and defence. Fibrous proteins form long, parallel strands with repetitive sequences; they are generally insoluble and suited to structural roles such as support and strength.

Haemoglobin: a globular protein

Haemoglobin has a quaternary structure made of four polypeptide chains: two alpha-globin chains and two beta-globin chains. Each chain holds one haem group, a prosthetic group containing one iron ion ($\text{Fe}^{2+}$) at its centre. So one haemoglobin molecule contains four haem groups and four iron ions.

Iron is the part that makes oxygen transport possible: it is the $\text{Fe}^{2+}$ ion in each haem group that the oxygen molecule actually binds to, and the iron must be in the iron(II) ($\text{Fe}^{2+}$) state for this binding to occur. Each iron ion binds reversibly to one oxygen molecule, so a single haemoglobin molecule can carry up to four oxygen molecules. Reversible binding is essential: oxygen is picked up where its concentration is high (the lungs) and released where it is low (respiring tissues). The chains fold so that the hydrophilic R-groups face outward, keeping haemoglobin soluble so it can be carried dissolved within red blood cells. The folding also positions each haem group correctly so oxygen can reach the iron.

Collagen: a fibrous protein

In short, the collagen hierarchy runs: three chains → one molecule (triple helix) → fibril → fibre.

A collagen molecule is made of three polypeptide chains wound tightly around one another to form a triple helix. Each chain is rich in the amino acid glycine, the smallest amino acid; because glycine has a tiny R-group, the three chains can pack very closely together. Many hydrogen bonds form between the chains, holding the triple helix firmly.

Many collagen molecules then line up alongside each other, staggered end to end, and are linked by covalent cross-links between the ends of neighbouring molecules. These bundles of molecules form fibrils, and many fibrils group together to form collagen fibres. The staggered arrangement and cross-links give the fibre great tensile strength.

Relating collagen structure to function

Collagen's structure makes it ideal for support and resisting pulling forces. The triple helix and the many hydrogen bonds make each molecule strong; the staggered overlap and covalent cross-links between molecules spread tension along the length of the fibre, so there are no weak points where the fibre would snap. This high tensile strength suits collagen to its roles in tendons (joining muscle to bone), skin, the walls of arteries and bone, where structures must withstand stretching without breaking.

Worked example

Exam-style question: Sickle-cell haemoglobin (HbS) differs from normal haemoglobin by a single change in the primary structure of its beta-globin chains. Explain how this single change can alter the behaviour of the whole protein. [3]

Model answer:

• The primary structure determines the position of the R-groups, so changing one amino acid places a different R-group at that point in the beta-globin chain.
• This alters the interactions (such as hydrophobic interactions) between R-groups, changing the tertiary/quaternary structure and so the surface shape of the haemoglobin molecule.
• At low oxygen levels the altered HbS molecules stick together (polymerise) into long fibres, which then distort the red blood cell into a sickle shape.

Worked example

Exam-style question: One molecule of haemoglobin is made of two alpha-globin chains and two beta-globin chains, each chain carrying one haem group. Each haem group contains one iron ion that binds one oxygen molecule. (a) Calculate the maximum number of oxygen molecules carried by one haemoglobin molecule. (b) A mutation changes one cysteine in a globin chain to alanine. Suggest how this could affect the protein. [4]

Model answer:

• (a) There are 4 chains, so 4 haem groups, so 4 iron ions; each iron ion binds 1 oxygen molecule, so the maximum is $4 \times 1 = \mathbf{4}$ oxygen molecules.
• (b) Cysteine R-groups form disulfide bonds, so losing a cysteine removes a disulfide bond from that point in the chain.
• With one fewer disulfide bond holding the fold, the tertiary structure of the chain may change shape.
• A changed shape can alter how the chains fit together or how the haem group sits, so the protein's function (oxygen binding) may be reduced or lost.

Key Equations

This topic is mainly qualitative. The only relationship needed is the formula for an amino acid and the condensation/hydrolysis of the peptide bond:

$$\text{amino acid + amino acid} \rightleftharpoons \text{dipeptide} + \text{H}_2\text{O}$$

The forward reaction (condensation) removes water to form the peptide bond; the reverse reaction (hydrolysis) adds water to break it.

Common Mistakes to Avoid

• Muddling the levels of collagen structure. Keep the order clear: three polypeptide chains twist together to make one collagen molecule (a triple helix); many molecules line up to form a fibril; many fibrils group into a fibre. Do not call a single molecule a "fibre" or "fibril".
• Treating a haemoglobin molecule and a red blood cell as the same thing. They are different levels: in sickle-cell anaemia it is the HbS molecules that polymerise and stick together, and this then changes the shape of the whole red blood cell.
• Saying haemoglobin has only one haem group. It has four haem groups (one per chain), each with one iron ion, so it can carry four oxygen molecules.
• Writing that the peptide bond forms by "adding water". A peptide bond forms by condensation (water removed); it is broken by hydrolysis (water added). Do not reverse these.
• Confusing which bonds hold which level. Hydrogen bonds maintain secondary structure (along the backbone) and also contribute to tertiary; hydrophobic interactions, ionic bonds and disulfide bonds act between R-groups in tertiary and quaternary structure.
• Saying the secondary-structure hydrogen bonds form between R-groups. In the secondary structure they form along the backbone, between the C=O of one peptide bond and the N–H of another; the R-groups only get involved later, in the tertiary structure.
• Calling the variable part of an amino acid the "carbon group". It is the R-group (side chain), and it is the only part that differs between amino acids.
• Saying oxygen binds to the protein chains directly. Oxygen binds to the iron ion in the haem group, not to the amino acids of the globin chains.

Exam Tips

• When asked to draw an amino acid, show the central carbon with all four attached groups clearly: $\text{-NH}_2$, $\text{-COOH}$, $\text{-H}$ and R.
• For "explain the structure of haemoglobin" questions, score the marks by naming two alpha chains, two beta chains, four haem groups and the iron ion, then linking each feature to oxygen transport.
• For collagen, always state the level you mean (chain, molecule, fibril or fibre) so your meaning is clear.
• Use "generally" when stating that globular proteins are soluble and fibrous proteins are insoluble, and always pair each type with its role (physiological vs structural).
• When asked to compare globular and fibrous proteins, use comparative connectives ("whereas", "in contrast") and pair both proteins in the same sentence for each point of comparison (shape, solubility, role, sequence), e.g. "globular proteins are roughly spherical and generally soluble, whereas fibrous proteins are long strands and generally insoluble". One-sided statements about only one protein do not score comparison marks.
• When a single amino acid change is mentioned, structure your answer as a chain of cause and effect: primary structure → R-group interactions → tertiary/quaternary shape → function.
• Spell the bond types precisely: disulfide bond (not "disulphate"), ionic bond, hydrogen bond and hydrophobic interaction.