7.7.2 Proteins

Primary Structure of Proteins

The primary structure of a protein refers to the specific sequence of amino acids within the polypeptide chain. This sequence is unique to each protein and is determined by the specific function the protein is intended to perform.

For example, a sequence might look like:

• gly – ala – leu – iso – gln (Each three-letter abbreviation represents a different amino acid.)

Proteins can be composed of thousands of amino acids, all arranged in a precise order to define the protein's properties. The primary structure is highly stable due to the strong covalent bonds (specifically, peptide bonds) that link the amino acids together in sequence.

Secondary Structure of Proteins

Protein chains do not remain straight, as hydrogen bonds can form between amino acids along the same chain or between neighboring chains. Specifically, hydrogen atoms on one peptide bond can interact with nitrogen or oxygen atoms on nearby peptide bonds, which can occur within the same molecule or between adjacent molecules.

This bonding results in two possible structural outcomes:

1. α-Helix The polypeptide chain coils into a helical structure, known as an α-helix. Typically, eighteen amino acids form approximately five coils in this structure.

β-Pleated Sheet

Alternatively, hydrogen bonds may primarily form between adjacent protein chains, creating a pleated sheet structure that spans 3 to 10 amino acids.

This localized three-dimensional arrangement of a protein, involving a small segment of adjacent amino acids, is called the secondary structure of the protein and typically takes the form of either an α-helix or a β-pleated sheet.

Tertiary Structure of Proteins

The tertiary structure of proteins refers to the interactions between amino acids that are located farther apart on the chain, which ultimately defines the protein's overall three-dimensional shape. There are three main types of interactions that stabilize and determine this tertiary structure:

2. Hydrogen Bonding Similar to the bonds that contribute to the secondary structure, hydrogen bonds form here between peptide links. A hydrogen atom attached to one peptide link can bond with nitrogen or oxygen atoms on another peptide link, helping to stabilize the protein's shape.

3. Ionic Bonding Some amino acids have an -NH₃⁺ group, while others contain a -COO⁻ group; these oppositely charged groups can attract each other, forming ionic bonds that contribute to the protein's tertiary structure.

4. Covalent Bonding (Disulfide Bonds) The amino acid cysteine contains a -CH₂SH group, which can interact with another cysteine's -CH₂SH group in the presence of oxygen to form a disulfide bond (S-S bond) as follows:

$$[-CH₂SH + -CH₂SH + [O] → -CH₂S-SCH₂- + H₂O]$$

These bonds form between amino acids that are distant from each other on the chain, influencing the protein's overall shape and thus contributing to its tertiary structure.

DNA (Deoxyribonucleic Acid)

DNA is the molecule that constitutes chromosomes, and thus genes, in all living organisms. Found within the cell nucleus, DNA is a polymer composed of repeating units known as nucleotides.

Nucleotides

A nucleotide is a molecule consisting of a phosphate group ($-H2PO4$) bonded to a deoxyribose sugar group ($-C5H8O2-$) which is turn bonded to a base group.

A nucleotide consists of:

• A phosphate group ($-H₂PO₄$),

• A deoxyribose sugar group ($-C₅H₈O₂-$),

• And one of four possible bases to which the sugar is bonded. The four different bases are:

• Cytosine (C)

• Thymine (T)

• Adenine (A)

• Guanine (G)

These bases define the four types of nucleotides, which are detailed in the AQA A-level Chemistry data sheet.

Polymerization of Nucleotides

DNA is composed of monomer units called nucleotides, which link together via condensation polymerization to form a single strand:

• In this process, an $OH$ group on the phosphate group of one nucleotide reacts with an $-OH$group on the sugar of another nucleotide, releasing water ($H₂O$).
• This reaction creates a sugar-phosphate backbone in which each sugar molecule is also bonded to one of four possible nitrogenous bases (adenine, cytosine, guanine, or thymine).
• This sequence forms one strand of DNA, which continues indefinitely to build a long chain.

Formation of the Double Helix

DNA exists as two complementary strands twisted into a double helix. Base pairing occurs between the strands:

• Adenine (A) pairs with thymine (T), and cytosine (C) pairs with guanine (G).
• These specific pairs form because the complementary shapes of the bases enable hydrogen bonding, stabilizing the double helix structure.

Hydrolysis of Peptides and Production of Amino Acids

Proteins are made up of amino acids joined by peptide bonds. When these peptide bonds are broken through hydrolysis, the protein decomposes into its individual amino acid components. This process requires the addition of water, typically in the presence of a strong acid or base, which cleaves the peptide link ($–CONH–$) between amino acids. This method allows for the identification of the original amino acids that constituted the protein.

• The $C-N$ bond in the peptide link is broken and the polypeptide/protein is broken down into its constituent amino acids in a hydrolysis reaction
• In the presence of water alone this process is very slow, but it can be catalysed by:
• An acid
• A substrate-specific enzyme.

Anti-Cancer Drugs: Cisplatin

Cisplatin is an effective anti-cancer drug used in chemotherapy, designed to target and disrupt the replication of cancer cells:

Mechanism of Action:

• Cisplatin binds to DNA by forming covalent bonds with two adjacent guanine bases on the DNA strand.

• This occurs through a ligand substitution reaction where the nitrogen atoms of guanine replace the chlorine atoms on cisplatin, resulting in covalent bonds between the platinum atom in cisplatin and the DNA bases.

• The bound cisplatin blocks DNA replication, as it prevents the DNA from unwinding and copying, thus halting cell division. Effectiveness and Side Effects:

• Cisplatin is effective because cancer cells divide rapidly, and the drug interferes more with these cells than with normal cells.

• However, it also affects healthy, rapidly dividing cells, like those in hair follicles, which explains why patients often experience hair loss.

• The use of cisplatin must weigh benefits against potential harmful side effects, with the goal being that the drug's ability to reduce cancer outweighs the impact on healthy cells. Design of Targeted Anti-Cancer Drugs:

• The most effective cancer treatments are those that selectively target cancer cells while minimizing damage to normal cells.

Separation and Identification of Amino Acids by Thin-Layer Chromatography (TLC)

• A mixture of amino acids formed by the hydrolysis of proteins can be separated and identified by TLC (thin-layer chromatography).
• The amino acids produced are colourless so they can only be located on a chromatogram by using developing agents such as ninhydrin (which stains the different amino acids) or UV light.
• The amino acids are then identified by measuring their Rf values. - Rf = distance moved by solute/distance moved by solvent - If you only have the chromatogram produced by the mixture of amino acids, you can compare the Rf value of each spot with data book values to identify the amino acid.
• A small sample of the amino acid mixture is applied to a stationary phase (usually a thin layer of silica gel or alumina on a glass or plastic plate).
• The plate is then placed in a solvent (mobile phase) that travels up the plate, separating amino acids based on their interactions with the stationary phase and their solubility in the solvent.

Visualization of Amino Acids on Chromatograms

Since amino acids are generally colourless, they need to be visualized using specific developing agents:

5. Ninhydrin: Reacts with amino acids to produce a purple or blue colour, making them visible on the chromatogram.
6. Ultraviolet (UV) Light: Some amino acids fluoresce under UV light and can be detected this way. After locating the amino acids on the chromatogram, their retention factor (Rf) values are calculated. The Rf value is specific to each amino acid and is calculated by the formula:

$$Rf= Distance travelled by the solvent front/ Distance travelled by the amino acid$$

These Rf values help in identifying the amino acids by comparing them to known standards.

Drawing the Structures of Peptides and Their Hydrolysis Products

1. Peptide Formation: A peptide is formed by linking two or more amino acids via condensation reactions, where the amino group ($–NH₂$) of one amino acid bonds with the carboxyl group ($–COOH$) of another, releasing water ($H₂O$).

• Example: A tripeptide could be drawn by linking three amino acids sequentially through peptide bonds.

2. Peptide Hydrolysis: When hydrolyzing a peptide, each peptide bond is broken, resulting in the release of individual amino acids. Diagrams should clearly show the individual amino acids with free amino ($–NH₂$) and carboxyl ($–COOH$) groups after hydrolysis.

Calculating Rf Values from a Chromatogram

To calculate the Rf value, you must measure the distance each amino acid spot has travelled from the baseline and divide it by the distance the solvent front travelled:

$$Rf = \frac{\text{Distance travelled by the amino acid}}{\text{Distance travelled by the solvent front}}$$

Rf values are a unique characteristic of each amino acid under specific conditions and can be used to identify amino acids by comparing with known Rf values.