B2 Proteins

B.2.1 Draw the general formula of 2-amino acids.

B.2.2 Describe the characteristic properties of 2-amino acids
Properties should include isoelectric point, formation of a zwitterion and buffer action.

Amino acids
Crystalline compounds with high melting points, usually above 200 degrees celsius
Greater solubility in water than non-polar solvents

Zwitterions - dipolar ions (positive and negative charge on same group of atoms)

Aminos are amphoteric, in Zwitterions it is the conjugate acid and base that's responsible for this property. In aqueous solution they can accept and donate H+ depending on the pH of the medium. As they can accept/donate it can resist small changes in pH - they can act as buffers, which is important to maintain body cell pH.
E.g. Enzymes are sensitive to pH change and can be destroyed easily by significant flucuations
pH determines net change
Positive (+) at low pH
Negative (-) at high pH

At the isoelectric point it is neutral at intermediate pH. There is no net change, amino acids will not move in an electric field, minimal repulsion, therefore least soluble.

B.2.3 Describe the condensation reaction of 2-amino acids to form polypeptides.
Reactions involving up to three amino acids will be assessed.
Different sequences = different peptides formed. (2 2-amino acids = dipeptide)
Uses condensation reaction - peptide and water are formed.

B.2.4 Describe and explain the primary, secondary (α-helix and β-pleated sheets), tertiary and quaternary structure of proteins. 
Include all bonds and interactions (both intramolecular and intermolecular) responsible for
the protein structure.

Primary structure = the amino acid sequence
Secondary structure = depends on the hydrogen bonding
Tertiary structure = depends on the interaction between R groups
Quaternary structure = association between different polypeptides

Primary structure (the sequence of amino acids)
- held together by peptide bonds
- dictates the entire structure and function of the protein
- extremely crucial, a change in one amino acid can change the entire functioning of the protein e.g. disease sickle-cell anemia. The disease occurs because of a single change in the chain of 146 amino acids, and the hemoglobin is unable to carry oxygen efficiently

Secondary structure (the folding of the polypeptide chain based on the side chains)
- the folding can differ in different proteins, or even in different sections of the same protein.
- the folding occurs as a result of hydrogen bonding between C=O and N-H groups of different peptide bonds
- 2 structures: α - helix & β - pleated sheet
- are fibrous proteins
- physically tough and insoluble in water

α - helix
- a tight coil, with 3.6 amino acids per turn, because of h-bonds formed between 2 peptide bonds 4 amino acids apart.
- flexible, elastic, because the intra-chain h-bonds break and reform easily as the molecule is stretched
- e.g Keratins, found in hair, skin + nails.

β - pleated sheet

- 'side by side' polypeptide
- extended form, not tightly wound
- pleated sheets cross-linked by inter-chain h-bonds
- flexible, inelastic
- e.g. fibers by spiders + silkworms, beaks

Tertiary structure (interactions between R groups)
Conformation
- relates to the most stable arrangement of the protein
- depends on the interaction of the R groups (the intra-molecular forces)
- important in globular proteins (this includes all enzymes and protein hormones)

Globular proteins 
- water soluble, because nearly all polar R groups are on the surface of the molecule they can interact with water, while the non-polar ones are mostly inside, out of contact with water.
The R-group interactions that stabilizes the conformation are the:
Hydrophobic interactions - between non-polar side chains
e.g. 2 alkyl side chains in valine, weak VDW between induced dipoles produce non-polar regions inside the protein.
Hydrogen bonding - between polar side chains
e.g. -CH2OH in serine and -CH2COOH in aspartic acid
Ionic bonding - between side chains carrying a charge
e.g. -(CH2)4NH3+ in lysine and -CH2COO- in aspartic acid
Disulphide bridges - between sulphur atoms in the amino acid cysteine
- covalent bonds, therefore the strongest interaction.

Interactions affected by changes in the medium e.g. temperature or pH
Denatured - when a protein loses its specific tertiary structure because of the changes
e.g. egg solidifying when heated, the enzymes denature - becoming biologically inactive
Therefore intra-cellular conditions must be controlled.


Quaternary structure (association between different polypeptide chains)
Only in some proteins, when they have more than 1 polypeptide chain.

e.g. Collagen (skin and tendons), most abundant protein in the human body
- triple helix of polypeptide chains
- inter-chain h-bonds, stable rope like formation that is resistant to stretching




e.g. Hemoglobin
- 4 polypeptide chains (α + β chains) fit together tightly

Most proteins consist of only 1 polypeptide, so they wouldn't have a quaternary structure




B.2.5 Explain how proteins can be analysed by chromatography and electrophoresis.
Analysis of proteins includes determining its amino acid composition - **NOT the primary structure, which is the sequence of amino acids**
It involves chemically separating amino acids (breaking peptide bonds through hydrolysis, usually with acid), reversing the condensation reaction.

Chromatography
- good to separate and identify mixtures (especially coloured ones e.g. felt pen ink)
- amino acids = colourless in solution but has colour when treated with a locating reagent (ninhydrin)

- small sample spotted at the bottom of chromatography paper (marked in pencil, the origin)
- paper suspended in a chromatographic tank with a small volume of solvent (where the spot is above level of solvent)

- solvent rises by capillary action, passes over the spot
- spot will distribute between 2 phases: stationary phase (water in the paper) and mobile phase (solvent)
- distributes at different rates, therefore will spread out according to their different solubilities
- solvent almost reaches the top and is marked (the solvent front)
- paper is removed and developed with ninhydrin (most amino acids turn purple, distinguished as separate isolated spots are up the length of paper
 Rf = retention factor

Electrophoresis
used to separate and identify intact proteins depending on their different rates of migration towards the electrodes
- analysis based on movement of charged particles in an electric field
- amino acids carry different charges depending on the pH and can be separated when placed in a buffer solution of a known pH (if pH is lower than their isoelectric point, the amino acid is positive)
- gel medium (usually polyacrylamide)
- amino acid mixture placed in wells in the center of the gel & electric field is applied
- depends on pH of buffer, different amino acids move at different rates towards oppositely charged electrodes
- at isoelectric points, amino acids will not move because they carry no net charge
- detected by a stain or floresence under UV light and identified by their position using data tables.

B.2.6 List the major functions of proteins in the body.
Include structural proteins (for example, collagen), enzymes, hormones (for example, insulin), immunoproteins (antibodies), transport proteins (for example, hemoglobin) and as an energy source.

Function of proteins