Protein Structure and Purification

Question 1

What configuration are amino acids in protein?

Answer 1

L-configuration

Question 2

What is the side chain for glycine?

Answer 2

H

Question 3

What is the side chain for alanine?

Answer 3

CH3

Question 4

What is the side chain for serine?

Answer 4

CH2OH

Question 5

What is the side chain for cysteine?

Answer 5

CH2SH

Question 6

How are peptide bonds formed?

Answer 6

Via a condensation reaction.

Question 7

What is the basic structure of an amino acid?

Answer 7

CHNH3RCOO

Question 8

Why is the C-N bond non-rotational?

Answer 8

Due to electron delocalisation, there is resonance.

Question 9

What is an amino acid residue?

Answer 9

The bit of the amino acid left behind after it joins a polpeptide chain.

Question 10

What are the two forms of secondary protein structure?

Answer 10

Alpha helices and beta sheets.

Question 11

What are some of the properties of alpha helices?

Answer 11

Right handed helix.
3.6 residues per turn.
Single helix.
H-bonds link across spirals to stabilise the helix.
Carbonyl of 1 links to amino 5.
n to n+4 linkage.
Side-chains/R groups project at 100 degrees.

Question 12

Which amino acid is a helix destabiliser?

Answer 12

Tyrosine (Y).

Question 13

Which amino acid is a helix breaker?

Answer 13

Proline (P).

Question 14

Why is proline a helix breaker?

Answer 14

(alpha)C-N bond cannot rotate meaning it has an exceptionally rigid structure and the R-group folds back on itself.

Question 15

What are the two forms of beta sheets?

Answer 15

Parallel and anti-parallel.

Question 16

How do beta sheets interact above and below?

Answer 16

They interdigitate - like fingers interlinking.

Question 17

Which form of beta sheets are stronger?

Answer 17

Anti-parallel beta sheets are stronger as the H-bonds are more linear.

Question 18

How does tertiary structure differ from secondary structure?

Answer 18

Tertiary structure involves side-chain interactions.

Question 19

Define hydrophobic collapse.

Answer 19

The idea that a protein would collapse rapidly around its hydrophobic side-chains and then rearrange from restricted conformational space occupied by the intermediate.

Question 20

Why is hydrophobic collapse favoured?

Answer 20

Hydrophobic collapse leads to greater disorder; positive entropy change.

Question 21

What are disulphide bonds?

Answer 21

Covalent bonds between two cysteine side-chains formed by oxidation.

Question 22

What is the Henderson-Hasselbach Equation?

Answer 22

pH = pKa + log(base/acid)

Question 23

What is a domain?

Answer 23

A globular unit formed from part of a polypeptide.

Question 24

What is a protein domain?

Answer 24

A conserved part of a given protein sequence that can evolve, function and exist independently.

Question 25

What is quarternary structure?

Answer 25

The assembly of more than one polypeptide chain.

Question 26

What are some examples of quarternary structure?

Answer 26

Haemoglobin and collagen.

Question 27

What is HIV proteinase?

Answer 27

A homodimer (two identical polpeptide chains) that helps produce the proteins for the virus, acting as a pair of scissors in a model kit.

Question 28

What is SDS-PAGE?

Answer 28

Sodium Dodecyl Sulphate - PolyAcrylamide Gel Electrophoresis. It is a type of protein purification using marker proteins of known molecular masses to estimate the molecular masses of other proteins.

Question 29

How does separation by solubility work?

Answer 29

Solubility of proteins differ because their surfaces differ.

Question 30

What is isoelectric focusing?

Answer 30

Utilises a protein's isoelectric point (the pH where a molecule has no net charge). Uses an immobilised pH gradient and an electric field. The proteins move towards the + or - end depending on their net charge. When they reach an area where their net charge is zero they stop moving.

Question 31

What is size exclusion chromatography?

Answer 31

Involves lots of tiny beads cross-linked with polydextran. Beads catch the smallest proteins as they can be 'pocketed'. Larger proteins get 'pocketed' less often.