SESSION 5

Question 1

What are the functions of proteins?

Answer 1

Proteins play crucial role in virtually all biochemical processes:

• catalysts
• transporters
• structural support
• muscular contraction and motion
• immune protection
• ion channels
• receptors
• ligands in cell signalling

Question 2

Describe the direct link between the DNA sequence and the protein sequence

Answer 2

DNA replication
Transcription
Translation
The nucleotide sequence of a gene determines the amino acid sequence of a protein

Question 3

What do amino acids consist of?

Answer 3

Amino acids consist of a central carbon atom covalently bonded to:

• an amino group (-NH2)
• a carboxyl group (-COOH)
• a hydrogen atom (-H)
• a distinctive R group (side chain)

Question 4

What are the ionised states of amino acids?

Answer 4

Both the carboxyl group and the amino group can ionise

BASE: NH2 + H+ NH3

ACID: COOH COO- + H+

Question 5

Define amino acid residue

Answer 5

An amino acid residue is what remains of an amino acid after it has been joined by a peptide bond to form a protein

Question 6

Classify amino acids by physical and chemical properties

Answer 6

Physical properties:

• aliphatic
• aromatic

Chemical properties:

• hydrophobic
• hydrophilic
• polar
• non- polar
• acidic
• base
• neutral

Question 7

Explain Pk values of ionisable side chains

Answer 7

An aid dissociation constant Ka is a quantitative measure of the strength of an acid in solution

Pk -> how likely is it to ionise

The lower the value, the more likely it is to be acidic, lose a proton

Question 8

Describe peptide formation

Answer 8

The linking of two amino acids is accompanied by the abstraction of a water molecule
Peptide bonds are created by enzyme catalysed condensation reactions
And broken down by enzyme catalysed hydrolysis reactions

Question 9

Describe the properties of peptide bonds

Answer 9

Peptide bonds are planar- all lie in the same place

Peptide bonds are rigid:

• partial double bond characteristics- less flexible and shorter
• unable to rotate

Bonds on either side of the peptide bond are free to rotate

Peptide bonds exhibit a tran’s conformation

Cannot form cis formation as it results in steric clashes

Question 10

Define conjugated proteins

Answer 10

Some proteins contain covalently linked chemical components in addition to amino acids

Question 11

What is the importance of amino acids in proteins?

Answer 11

The amino acid sequence of a protein determines:

• the way in which the polypeptide chain folds
• the physical characteristics of the protein

Question 12

Define the isoelectric point

Answer 12

The isoelectric point of a protein is the pH at which there is no overall net charge

PI > 7 - contain many positively charged amino acids

PI < 7 - contain many negatively charged amino acids

If the pH of the solution < the pK value then the group will be protonated
If the pH of the solution > the pK value then the group will be deprotonated

Question 13

Define peptide

Answer 13

A few amino acids in length

Question 14

Define polypeptide

Answer 14

Many amino acids

Question 15

Describe what is meant by primary, secondary, tertiary and quaternary structure of proteins

Answer 15

Primary structure- the amino acid sequence of a protein

Secondary structure-
stretches of the polypeptide chain that form a-helical and B- sheets.
Bonds on either side of the peptide bond can rotate freely
When these angles remain the same throughout a segment of polypeptide the protein adopts a regular secondary structure

Tertiary structure-
The full 3D structure of the protein 
Involves the folding up of the secondary structures
Improper folding (amyloidosis) may cause disease
Most proteins fold spontaneously, but some require help of molecular chaperones 

Quaternary structure-
Interaction between and arrangement of different polypeptide chains (subunits) within the same protein
The polypeptide chains may be identical (homomeric) or different (heteromeric)

Question 16

Define both globular proteins and fibrous and their roles

Answer 16

Globular- 
several types of 2 structure
Compact shape 
Role: Enzyme and regulatory proteins 
E.g. Carbonic anhydride 

Fibrous
Single type of repeating secondary structure
Long strands or sheets
Role: Structure, support, protection

Question 17

Describe the type of bonds and forces involved in protein structure

Answer 17

Primary structure- covalent bonds (peptide)

Secondary structure - hydrogen bonds

Tertiary structure- hydrogen bonds/ van der waals/hydrophobic interactions/ covalent (disulphide)/ ionic interaction

Quaternary structure- same as tertiary

Question 18

Explain the key features of the two major secondary structure elements of proteins (a-helix and B- sheets)

Answer 18

B-sheet:

• extended conformation
• parallel or antiparallel
• multiple inter- strand H- bonds
• e.g. Fatty acid binding protein

A- helix:

• right- handed helix
• 3.6 amino acids per turn
• small hydrophobic AA’s such as Ala + Leu are strong helix formers
• pro acts as a helix breaker because the rotation around N- Ca bond is impossible
• Gly acts as a helix breaker because the tiny R- group supports other conformations. E.g. Iron storage protein ferritin

Question 19

Explain the physiological roles of myoglobin and haemoglobin

Answer 19

Myoglobin-
a single subunit protein that contains one haem group for the binding and transport of oxygen. It can bind one molecule of oxygen. Mb exhibits hyperbolic O2 binding - no cooperativity

Haemoglobin-
A tetrameric protein (2xa, 2xB subunits) containing four haem groups.
It can bind our molecules of oxygen
Hb exhibits sigmoidal O2 biding

Question 20

Contrast the oxygen- binding properties of myoglobin and haemoglobin and explain why haemoglobin is most suited to its role of an oxygen transporter

Answer 20

Mb has a very high affinity for O2, thus will only release O2 when pO2 is extremely low

Hb however can exist in two states- a low affinity T- state and a high affinity R- state
Transition between these two states gives Hb it’s sigmoidal binding curve- Hb’s affinity to O2 increases as more O2 binds

Question 21

Describe the major structural differences between oxygenated ad deoxygenated haemoglobin and the molecular basis of cooperativity

Answer 21

Deoxyhaemoglobin can exist in a low affinity T-state or a high affinity R state
The binding of O2 promotes the stabilisation of the R state- cooperatively

Question 22

Describe the effects of CO2, H+ and 2-3- bisphosphoglycerate and CO on the binding of O2 by haemoglobin, and the physiological significance of these effects

Answer 22

CO2 and H+ decrease the affinity of Hb for O2. At sites of low pH and increased CO2, e.g. Muscle tissue during exercise, more oxygen is required and will be released. This is called the Bohr effect

2-3- bisphosphoglycerate (BPG) decreases the affinity of Hb four O2. Curve shifts right
BPG concentration increases at high altitudes, promoting O2 release at tissues.
BPG is also produces during metabolism, so O2 is released more readily in areas performing high amounts of metabolism

CO binds to Hb 250X more readily than oxygen. It is fatal when COHb is > 50%. The binding of CO also acts to increase the affinity of unaffected subunits for oxygen

Question 23

Appreciate that mutations in Globin can give rise to diseases such as sickle cell anaemia or thalasaemia

Answer 23

Sickle cell anaemia
SSA is an autosomal recessive genetic blood disorder.
It is caused by a single nucleotide polymorphism resulting in the substitution of (hydrophilic) Glutamate -> Valine (hydrophobic) in the B- subunit of Hb
A sticky hydrophobic pocket formed by Val allows deoxygenated Hbs to polymerise
This leads to the distortion of RBCs into the sickle cell shape.
This distortion causes stress to the cell membrane and causes premature cell lysis
The sickle cell also blacks microvasculature

Thalassaemias
Thalassamias are a group of genetic disorders where there is an imbalance between a and B subunits
B- thalassamias- decreased or absent B- chains. A- chains are noble to form stable tetrameters. Symptoms appear after birth
A- thalassamias- decreased or absent a- chains. There are several different levels of severity as there are multiple copies of the a- chain gene present. B- chains can form stable tetrameters with increased affinity for oxygen. Onset before birth

Question 24

Explain why some amino acid side chains are charged at physiological pH

Answer 24

If the pH of the solution < the pK value then the group will be protonated
If the pH of the solution > the pK value then the group will be deprotonated

Question 25

What does the pKa value of an amino acid side chain tell you about that chemical group?

Answer 25

PKa value - log of the acid dissociation constant
Negatively charged R groups have a lower pK value
Positively charged R groups have a higher pK value

The lower the pK value the more likely it is for the R group to loose a proton and be in an acidic form
The higher the pK value the more likely it is for something to be basic

Question 26

Why do you think histones have a positive charge?

Answer 26

DNA backbone is negatively charged due to the presence of phosphate groups and this is to allow tight association between the histone and he DNA backbone in order to control packaging and transcription