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ESA 1 - Medical Cell Biology & Genetics > Session 5.2a - Lecture 1 - Protein Folding and Protein Structure Review > Flashcards

Session 5.2a - Lecture 1 - Protein Folding and Protein Structure Review Flashcards

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1
Q

ILO

A

Review of protein structure

  • Protein structure review
  • Enzymes review (Session 6)
  • AAs that make up proteins and their properties

Protein folding

  • How do proteins actually fold
  • How do they get their 3D shape
  • Important interactions that help protein folding occur
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2
Q

Name the 4 layers of protein structure.

A

Primary structure
Secondary structure
Tertiary structure
Quaternary structure

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3
Q

Explain how the first 2 layers of protein structure are related.

A

The primary sequence folds to give a localised secondary sequence.

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4
Q

Give 2 examples of secondary structure conformations.

A
  • Alpha helix

- Beta sheet or beta strands

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5
Q

What is the structure of alpha helix secondary structure?

A
  • Helical shape
  • Relatively compact
  • H-bonds running up and down chain to stabilise it
  • R groups on outside
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6
Q

What shape is the alpha helix secondary structure?

A

Helical and relatively compact

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7
Q

Where do the R groups lie in the alpha helix secondary structure?

A

On the outside

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8
Q

What is the structure of beta sheet/strand secondary structure?

A
  • More extended conformation

- H bonds stabilise it

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9
Q

What stabilises the alpha helix and beta sheet secondary structures?

A

H-bonds

  • running up and down chain in alpha helix
  • between individual strands in beta strands
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10
Q

What is the tertiary structure of a protein?

A

Where parts of the molecule fold up, so parts that are from far apart in the original polypeptide sequence may be close together

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11
Q

How are some amino acids that are far apart in the original polypeptide sequence found close together in proteins?

A

Due to folding of the tertiary structure

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12
Q

What element of proteins do only some proteins contain?

A

Quaternary structure

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13
Q

What is quaternary structure?

A

Where there is distinct subunits - >1 subunit but we can also it could be interactions with macromolecules (big molecules, e.g. DNA, RNA) in other constituent elements.

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14
Q

Give 2 examples of macromolecules found in the human body.

A

DNA

RNA

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15
Q

Does myoglobin have quarternary structure?

A

No, it is monomeric, has 1 single polypeptide chain

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16
Q

Myoglobin can bind to a haem group. Does this mean it has quarternary structure?

A

No, because haem isn’t really a macromolecule so we don’t take that into account.

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17
Q

Give an example of a protein with quarternary structure.

A

Haemoglobin - made up of 4 subunits.

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18
Q

How many subunits is haemoglobin made up of?

A

4

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19
Q

Fig. 2

Label this image

A

Protein structure and function

Primary structure
Secondary structure
Tertiary structure
Quarternary structure

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20
Q

Draw the different layers of protein structure.

A

See Fig. 2

Protein structure and function

Primary structure
Secondary structure
Tertiary structure
Quarternary structure

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21
Q

What are the forces involved in maintaining protein structure in the primary sequence?

A

Covalent (peptide)

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22
Q

What are the forces involved in maintaining protein structure in the secondary sequence?

A

H-bonds

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23
Q

What are the forces involved in maintaining protein structure in the tertiary sequence?

A
  • Covalent (disulphide)
  • Ionic
  • H-bonds
  • van der Waals
  • Hydrophobic
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24
Q

What are the forces involved in maintaining protein structure in the quaternary sequence?

A
  • Covalent (disulphide)
  • Ionic
  • H-bonds
  • van der Waals
  • Hydrophobic
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25
Q

How are covalent bonds involved in protein structure?

A

Primary structure (peptide)

Tertiary and Quaternary structure (disulphide)

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26
Q

How are hydrogen bonds involved in protein structure?

A

Secondary,
Tertiary &
Quaternary structure

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27
Q

Explain the forces we see in primary structure.

A

In primary – only covalent bonds involved, i.e. peptide bonds – only thing that holds individual AARs together

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28
Q

Where are peptide bonds found?

A

Between amino acid residues in the primary structure of a protein.

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29
Q

Explain the forces we see in secondary structure.

A

A key element to see in secondary structure is that ALL of it has formed by H-bonds, so if we remember alpha helices: H-bonds between carbonyl-oxygens and amide-hydrogens running up the chain. In our beta strands, H-bonds between individual strands

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30
Q

Give an example of a covalent bond found in tertiary and/or quaternary structure.

A

Disulphide

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31
Q

Why are vdW interactions important in tertiary and/or quaternary structure?

A

Important particularly to think about vdW interactions, relatively weak but characteristic of any covalent bond, so although incredibly weak, think of a protein made up of 100s of AA residues, each of which contains several 10s of covalent bonds can see vdW becomes increasingly important.

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32
Q

What is the strongest type of bond in the protein?

A

Covalent bond

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33
Q

What types of covalent bonds can you find in proteins?

A

Peptide bonds (between amino acids)

Disulphide bonds

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34
Q

What are disulphide bonds?

A

Covalent bonds formed between 2 cysteine residues.

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35
Q

Cysteine is able to form what?

A

Disulphide bonds

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36
Q

Why can cysteine form disulphide bonds?

A

They have a sulphydryl group (-SH) at the end of their R group.

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37
Q

What is a disulphide bond?

A

A covalent bond formed between 2 sulfur atoms.

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38
Q

What type of reaction forms a disulphide bond?

A

Oxidation reaction

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39
Q

Write the reaction that forms a disulphide bond between 2 sulphydryl groups.

A

SH + SH

– 2H+ + 2e- –>

S-S

(reaction is reversible)

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40
Q

How can you break down disulphide bonds?

A

Can be broken down by adding reducing agents: that can become important in some conditions

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41
Q

What is clinically important about disulphide bonds?

A

Can be broken down by adding reducing agents, which is important in some conditions

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42
Q

Give an example of a reducing agent that breaks down disulphide bonds.

A

b-mercaptoethanol

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43
Q

What is b-mercaptoethanol?Name 3 facts.

A

A reducing agent that can break down disulphide bonds.

It is used in SDS-PAGE gels

Smells strongly of sulphur.

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44
Q

Why do we need disulphide bonds in proteins?

A

These are mainly for proteins that get secreted.

Inside your cells we have quite a stable environ, we can control what sort of elements or conditions that protein see – proteins actually fall apart quite easily as and when needed.

However, outside the cell there is a hostile environment, so diS bonds help maintain protein structure for stability - it acts as a protective mechanism

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45
Q

In which proteins do we tend to find disulphide bonds?

A

Tend not to see them in many proteins inside your cells but become v important for proteins that get secreted

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46
Q

Why don’t we see disulphide bonds in proteins inside the cell?

A

Inside your cells we have quite a stable environ, we can control what sort of elements or conditions that protein sees – in fact, proteins actually fall apart quite easily. Thus, they are not needed here.

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47
Q

Why do we need disulphide bonds for proteins going to the lumen of the gut?

A

Proteins that get secreted to the lumen of your gut are going to a much more hostile environment, compared to the safe interior of the cell.

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48
Q

Give an example of an enzyme that has disulphide bonds in its structure.

A

Ribonuclease enzyme

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49
Q

What is the function of ribonuclease?

A

Chop up RNA.

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50
Q

Why does ribonuclease need disulphide bonds?

A

The ribonuclease enzyme is involved in chopping up RNA that you might find, in your gut, for example.

This is a hostile environment, thus the disulphide bonds prevent the enzyme from breaking down.

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51
Q

How much energy does it take to break a disulphide bond?

A

214 kJ/mol

strong

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52
Q

Fig. 4 (right)

Label this image.

A

Cysteine SH
Cysteine SH

2H+ + 2e- x2

Cystine S-S

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53
Q

Fig. 4 (left)

Caption and label this protein.

A

Most proteins with disulphide bonds are secreted

e.g. ribonuclease

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54
Q

Draw the reaction of two cysteines forming a disulphide bond.

A

See Fig. 4 (right)

Cysteine COO–NH3+-CH-CH2-SH
SH-CH2-CH-NH3+-COO- Cysteine

– 2H+ + 2e- –>

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55
Q

Draw the structure of a ribonuclease enzyme

A

See Fig. 4 (left)

spiral - include 4 cysteine bonds that hold tertiary structure together

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56
Q

Give some facts about covalent (disulphide) bonds.

A
  • Formed between Cys residues
  • 214 kJ/mol
  • Can be broken by with reducing agents
    e. g. b-mercaptoethanol
  • Most proteins with disulphide bonds are secreted
    e. g. ribonuclease
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57
Q

What are the forces involved in maintaining protein structure?

A
  • Covalent (disulphide) bonds
  • Electrostatic interactions
  • Hydrogen bonds
  • Hydrophobic effect
  • van der Waals forces
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58
Q

What produces electrostatic interactions in proteins?

A

Formed between charged groups

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59
Q

Give examples of groups which can form electrostatic interactions in proteins

A

e.g. Glu-, Asp- and Arg+, Lys+, His+ (honorary member)

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60
Q

Why is histidine different to the other basic amino acids?

A

It is weakly charged, in fact, at physiological pH most of His is not negatively charged - however it gets put into this group.

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61
Q

How strong are electrostatic interactions?

A

10-30 kJ/mol

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62
Q

How often do we see H bonds in proteins?

A

Often

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63
Q

What forms a hydrogen bond?

A

Formed between electronegative atom and a hydrogen bound to another electronegative atom

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64
Q

Define hydrogen bond.

A

Formed between electronegative atom and a hydrogen bound to another electronegative atom - strictly speaking, these must be an oxygen, nitrogen or fluorine.

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65
Q

What atoms are hydrogen bonds in proteins formed by?

A

By definition - must be H and either O, N or F, but in proteins we are mainly thinking about interactions between (H and) N and O, can probably forget F.

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66
Q

What sort of groups can form hydrogen bonds?

A

Lots of different groups!

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67
Q

How strong are hydrogen bonds?

A

10-30 kJ/mol

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68
Q

What are electrostatic interactions in a protein sometimes called?

A

Salt bridge

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69
Q

What is a salt bridge?

A

Electrostatic interactions in a protein between charged groups.

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70
Q

Give some facts about electrostatic interactions.

A
  • Formed between charged groups
    e. g. Glu-, Asp- and Arg+, Lys+, His+
  • 10-30 kJ/mol
  • (Salt bridge)
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71
Q

Give some facts about hydrogen bonds.

A
  • Formed between electronegative atom and a hydrogen bond to another electronegative atom
  • 10-30 kJ/mol
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72
Q

Fig. 5

Label this image.

A

Hydrogen acceptor

Hydrogen donor

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73
Q

Draw some different examples of H-bonds.

A

See Fig. 5

Hydrogen acceptor
Hydrogen donor

  • C=O…H-O
  • N…H-O
  • O…H-O
  • C=O…H-N
  • O…H-N
  • N…H-N
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74
Q

What is the hydrophobic effect?

A

Interaction between hydrophobic side chains

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75
Q

Why does the hydrophobic effect occur?

A

Due to the displacement of water

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76
Q

What is the hydrophobic effect?

A

The interaction of hydrophobic side chains driven by the desire to exclude water.

It is not an interaction like electrostatic interactions, however - they are ‘forced’ together.

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77
Q

Why is the hydrophobic effect not the same as hydrophobic bonds?

A

Hydrophobic effect is interaction between hydrophobic side chains – they’re not effectively interacting together like electrostatic interactions but they’re kind of forced together by the exclusion of water (this drives formation of these interactions).

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78
Q

How strong are hydrophobic effect interactions?

A

~10 kJ/mol

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79
Q

What are van der Waals forces?

A

Dipole-dipole interactions

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80
Q

Where are vdW forces found?

A

Between any covalent bond - at any one time, there will be a slight variation of distribution of electrons

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81
Q

When are vdW forces important?

A

Important in big molecules like proteins that are coming together, or when surfaces of 2 large molecules come together

82
Q

How strong are vdW forces?

A

4 kJ/mol

weak

83
Q

Name some facts about the hydrophobic effect.

A
  • Interaction between hydrophobic side chains
  • Due to displacement of water
  • ~10 kJ/mol
84
Q

Name some facts about van der Waals forces.

A
  • Dipole-dipole interactions
  • 4 kJ/mol
  • Important when surfaces of 2 large molecules come together.
85
Q

Fig. 6

Label the image

A

Nonpolar molecule x4

86
Q

Draw an image depicting the hydrophobic effect.

A

See Fig. 6

Nonpolar molecule x4

87
Q

List the forces involved in maintaining protein structure from strong to weak, with their associated strengths.

A
  • Covalent (disulphide) bonds = 214 kJ/mol
  • Electrostatic interactions = 10-30 kJ/mol
  • Hydrogen bonds = 10-30 kJ/mol
  • Hydrophobic effect = ~10 kJ/mol
  • van der Waals forces = 4 kJ/mol
88
Q

Name the interactions involved in the forces involved in maintaining protein structure

A
  • Covalent (disulphide) bonds = 2 -SH groups (cysteine) make S-S
  • Electrostatic interactions (salt bridge) = between charged groups (Arg+ Lys+ His+ Glu- Asp-)
  • Hydrogen bonds = H and N O F (electronegative atom)
  • Hydrophobic effect = hydrophobic side chains driven together by exclusion of water
  • van der Waals forces = dipole-dipole interactions in covalent bonds (partial charges)
89
Q

How easy is protein denaturation?

A

Proteins fall apart very easily - think PhD scientist keeps losing their work bc their protein keeps falling apart!

90
Q

Why do proteins fall apart so easily?

A. To get on the PhD scientists nerves
B. Because they suck
C. Other - explain.

A

C. Proteins are not very stable

91
Q

Where are proteins likely to fall apart?

A

Proteins once outside realm of cell will fall apart relatively easily

92
Q

What do we mean when we talk about a normal protein?

A

A normally folded protein that is functional is said to in the NATIVE conformation

93
Q

What is the native conformation of a protein?

A

A normally folded protein that is functional is said to in the NATIVE conformation

94
Q

What is denaturation?

A

A normally folded protein that is functional is said to in the NATIVE conformation

Disruption of protein structure is known as denaturation - i.e. moving them from their native state

95
Q

How is a protein denatured?

A

Can be done in lots of different ways, but the underlying mechanism involved breaking of forces that hold proteins together.

96
Q

Give 3 examples of ways you can denature a protein.

A
  • Heat
  • pH (extremes of, either high or low)
  • Detergents/organic solvents
97
Q

Why does heat denature a protein?

A

Increased vibrational energy

98
Q

Give a classic example of how heat denatures a protein.

A

Classic example is frying an egg – proteins in an egg are originally liquid, they then become solid due to denaturation - adding heat thus actually loses the original shape.

99
Q

How does pH denature a protein?

A
  • Alters ionisation states of amino acids

- Changes ionic/H-bonds

100
Q

What is the effect of changing the charge on amino acid residue side chains, and how could that occur?

A

It would change the electrostatic, ionic and H-bonds in a protein, thus denature it.

101
Q

How can changing the ionisation state of amino acids be effected?

A

This can occur by changing the pH to extremes (either high or low)

102
Q

What do adding detergents/organic solvents do to a protein?

A

Disrupt hydrophobic interactions

103
Q

How can we disrupt hydrophobic interactions?

A

By adding detergents/organic solvents to a protein

104
Q

Why is it clinically important to know that proteins can fall apart easily?

A

. So proteins can fall apart v easily but we can also exploit the use of that in our studies when we start to think about molecular diagnosis.

105
Q

Explain what protein denaturation is and how that can occur

A

Protein denaturation

  • Proteins are not very stable
  • A normally folded protein that is functional is said to in the NATIVE conformation
  • Disruption of protein structure is known as denaturation
  • Caused by breaking of forces that hold proteins together
    e. g. - Heat: Increased vibrational energy
  • pH: Alters ionization states of amino acids; Changes ionic/H-bonds
  • Detergents/Organic solvents: Disrupt hydrophobic interactions
106
Q

Where does a protein shape come from?

A

They start with their ‘beads on a string’ conformation - completely undefined state (amino acid sequence) and fold into their 3D conformation.

107
Q

Why is it important that proteins are in a 3D conformation?

A

They’re only going to be active, only going to do what we want them to do when they’re in the right shape – if not, they’re not really going to work

108
Q

How do proteins fold?

A
  • All the information needed for folding is contained in the primary sequence
109
Q

How do we know that the primary sequence contains the information for protein folding?

A

Can be seen from a lab experiments:

Take a folded protein and you measure its activity in some way, e.g. an enzyme, can measure quite easily – then add a denaturing agent such as urea. This abolishes all tertiary and secondary structure. So you get this sort of unwound polypeptide sequence, and what you see is this activity eventually reaches 0 – so it is fully unfolded. If you take the urea away, so you can dialyse this away to reduce the urea concentration, what you can see for many proteins is they will go back and refold, and regain their activity. I’ve put nothing else in the test tube – so enzyme has denatured, renatured and maintained activity again. So the only info that this protein actually has is contained within its primary sequence.

110
Q

Give an example of a protein where you can measure its activity.

A

An enzyme

111
Q

Give an example of a denaturing agent.

A

Urea

112
Q

If you add a denaturing agent, such as urea, to an enzyme - how can you remove the urea from the preparation?

A

By dialysing it to reduce the urea concentration

113
Q

What happens to enzyme activity if you add a denaturing agent and then take that away again?

A
  • Enzyme activity begins at 100% with no denaturing agent
  • Increased concentration of urea reduces enzyme activity until it eventually reaches 0%
  • As you take the urea away again the concentration increases inversely proportional to the urea concentration until all urea is gone/enzyme activity reaches 100% again
114
Q

What is the relationship between enzyme activity and viscocity?

A

It is inversely proportional: as enzyme activity decreases the viscocity increases

This shows the change from maximal effect at 3D conformation, to loss of tertiary shape and unwounded primary sequence polypeptide (100% viscosity, 0% activity)

115
Q

What is the function of the primary sequence of the protein?

A
  • Holds the unique amino acid sequence of the protein
  • This gives it its chemical and physical properties of the proteins
  • It also gives the information for the 3D structure of the protein, necessary for function (structure related to function)
116
Q

Fig. 8

Label this graph

A

activity (%) 100 75 50 25 0
urea concentration (M) 0 8 0
viscosity (&) 100 75 50 25 0

117
Q

Draw a graph depicting the enzyme activity of a protein in relation to its shape.

A

See Fig. 8

activity (%) 100 75 50 25 0
urea concentration (M) 0 8 0
viscosity (&) 100 75 50 25 0

include drawing of 3D native conformation and single polypeptide denatured conformation in accordance with activity/viscosity.

118
Q

Explain what would happen if protein folding is random.

A

Protein folding cannot be random – would take too long!

119
Q

Describe the size of most proteins and relate it to protein folding.

A

Most proteins are fairly large.

Even a 100 AA protein, which is really small compared to a lot of proteins, would take a lot of time (billions of years!) to fold if protein folding was random!

Thus protein folding cannot be random – would take too long!

120
Q

Explain why protein folding cannot be random with the example of a 100 amino acid protein.

A

Protein folding cannot be random – would take too long!

100 amino acid protein

  • at least 3 possible values for each phi and psi bond
  • 3^198 different conformations
  • assuming that the protein could explore 10^13 conformations per second

This would take 3 x 10^80 years!!!!

121
Q

How many bonds does each amino acid have?

A

2 - a phi and psi bond

122
Q

How many conformations can each phi and psi bond have?

A

At least 3 possible values for each phi and psi bond (actually quite a low number of conformations)

123
Q

How many different conformations would a 100 amino acid protein be able to make?

A
  • Each amino acid has 2 bonds (phi and psi bonds)
  • Each of these has 3 possible conformations
  • The end two amino acids only have 1 bond which can rotate
  • so 100 amino acids x 2 bonds = 200
  • 200 bonds - 2 bonds = 198

-3^198 different conformations

124
Q

How quick can proteins make conformations?

A
  • assuming that the protein could explore 10^13 conformations per second
125
Q

Assuming a protein can explore 10^13 conformations per second, how long would it take a 100 amino acid protein to fold randomly?

A

This would take 3 x 10^80 years!!!!

126
Q

Assuming a protein can explore 10^13 conformations per second, why is it impossible for proteins to fold randomly?

A

This would take 3 x 10^80 years!!!! (3^198 different possible conformations)

This cannot occur in humans!

127
Q

What is the mechanism behind protein folding, random chance or other?

A

It can’t do it just by random chance as it would take far too long – so there must be some sort of mechanism to drive random folding of proteins.

128
Q

Describe the process by which proteins fold.

A

The folding process must be ordered

Each step involves localised folding and with stable conformations maintained

129
Q

Explain whether protein folding is random or ordered.

A

Protein folding cannot be random because it would take too long with all the possible conformations!

So what we tend to see is that proteins go through a semi-ordered structure.

130
Q

What do we mean by semi-ordered protein folding?

A

So they start off in a random conformation, and then by chance they get some sort of localised folding, and at some point part of the localised folding will by chance be correct. Once they’ve adopted the right state, they’ll lock into that state - which significantly reduces the number of conformations possible, thus the time it would take to do this.

131
Q

Give an analogy to the mechanism of protein folding.

A

e.g. a random typing of Shakespeare would take an infinite (or at least very large!!) amount of time but if you maintain keystrokes that are correct then this is much faster

132
Q

What do we mean by “stable conformations maintained” when describing protein folding?

A

Things start off in a random conformation and undergo localised folding by chance.

So actually we’ve got this idea of localised folding – get things locking in to be held in this state.

133
Q

Fig. 10

Explain this image.

A

e.g. a random typing of Shakespeare would take an infinite (or at least very large!!) amount of time but if you maintain keystrokes that are correct then this is much faster

134
Q

Draw an image explaining how proteins fold.

A

See Fig. 10

e.g. a random typing of Shakespeare would take an infinite (or at least very large!!) amount of time but if you maintain keystrokes that are correct then this is much faster

135
Q

Explain how proteins fold.

A

Proteins cannot fold randomly, bc this would take too long - at a rate of 10^13 conformations per second, and 3 conformations per phi or psi bond - a 100 AA protein would take 3 x 10^80 years! Clearly this is impossible in humans, so protein folding cannot be random.

What we actually find, it it is SEMI-random/ordered. Proteins start off in a completely random conformation, and by chance adopt a conformation through localised folding. This conformation can be either ‘wrong’ or ‘right’ - once the protein has found the correct conformation - it locks this conformation in place. This significantly reduces the number of conformations possible and thus time taken, until the whole protein is locked in the right place.

An example is using the Shakespeare monkey analogy. It would take an infinite long number of time for them to type out a Shakespearean sonnet randomly. However, if every time they randomly get a letter write, and ‘lock’ it in place - this would take about 2883 tries to get 1 correct sentence: thus, reducing the time from infinite to manageable.

136
Q

Describe simply the mechanism of protein folding.

A

It is a STEP-WISE process - where each correct conformation is locked in.

137
Q

What drives protein folding?

A

Entropy - driven by the need to find the most stable conformation

138
Q

If protein folding occurs in a step-wise fashion, what would this create?

A

Partially folded intermediates

139
Q

How are partially folded intermediates formed?

A

Due to the creation of protein folding - as it doesn’t happen all immediately but in a step-wise fashion.

140
Q

What are partially folded intermediates of proteins called?

A

Molten globule state

141
Q

What is the molten globule state?

A

A partially folded intermediate

142
Q

What is the sequence of protein folding?

A

Unfolded polypeptide –> partially folded intermediates –> fully folded protein

143
Q

Do most proteins fold independently?

A

Most proteins can do this completely on their own – as proteins are being made on ribosomes they will start to fold. All the info they’ve got and need to fold is contained in primary sequence.

That doesn’t mean some proteins don’t need a bit of help – some need to be helped by things called molecular chaperones

144
Q

Some proteins need a bit of help to fold. What helps these proteins?

A

Molecular chaperones

145
Q

What are molecular chaperones?

A

Proteins that bind to partially unfolded shapes to help stabilise them, stopping bits of protein we don’t want to come together coming together, which would otherwise cause an abnormal sequence.

146
Q

What might happen if protein folding goes wrong?

A

The structure of a protein defines what it does. Thus, if protein folding went wrong, then we might end up with a diff structure, which then might not do what we want it to do.

147
Q

Explain how proteins fold.

A
  • The folding process must be ordered
  • Each step involves localised folding and with stable conformations maintained
  • Driven by the need to find the most stable conformation
148
Q

Fig. 11

Label this image

A

unfolded polypeptide –> partially folded intermediates –> fully folded protein

149
Q

Draw an image depicting protein folding.

A

See Fig. 11

unfolded polypeptide –> partially folded intermediates –> fully folded protein

(starts with single polypeptide then adds secondary structures (alpha helix/beta sheets) until these are put into tertiary structure of 3D shape)

150
Q

Why is protein misfolding clinically important?

A

Protein misfolding can cause disease

151
Q

What group of diseases is often caused by protein misfolding?

A

Transmissible spongiform encephalopathies

152
Q

What are transmissible spongiform encephalopathies (TSEs)?

A

TSEs are a group of progressive, invariably fatal, conditions that are associated with prions and affect the brain (encephalopathies) and nervous system of many animals, including humans, cattle, and sheep

153
Q

Give 3 examples of TSEs.

A

Transmissible spongiform encephalopathies

  • Bovine spongiform encephalopathy (BSE)
  • Kuru
  • Creutzfeldt-Jakob disease (CJD)
154
Q

How can protein misfolding cause disease?

A

Altered conformation of a normal human protein promotes converts existing protein into diseased state

155
Q

What is a prion?

A

Prions are misfolded proteins which characterize several fatal neurodegenerative diseases in humans and many other animals (TSEs, e.g. BSE, kuru, CJD)

156
Q

How do TSEs arise?

A

These involve these things known as prion state – what these involve are abnormal folding of normal constituent human proteins in many cases (the prion protein - PrP)

157
Q

What protein is affected in prions?

A

The PrP protein

158
Q

What is the normal form of the prion protein?

A

PrP^C (c = cellular)

159
Q

What is the infectious form of the prion protein?

A

PrP^Sc (sc = scrapie)

160
Q

What is the PrP protein?

A

It is a normal neuronal protein that is found throughout the body in healthy people (and animals), but it is the protein that prions are made of. In its normal form, it is PrP^C, whereas the infectious form is known as PrP^Sc. The two have different conformations, alpha helical and beta sheet/strands respectively.

161
Q

What is the conformation of PrP^C?

A

PrP^C is the normal neuronal state of the protein - in its normal native state has quite a lot of alpha helices.

162
Q

What is the conformation of PrP^Sc?

A

PrP^SC is the disease state of the prion protein - in its disease state has quite a lot of beta sheets (strand-like structures).

163
Q

Describe the two states of the PrP protein.

A

PrP^C (normal) = lots of alpha-helices

PrP^Sc (disease) = lots of beta sheets/strands

164
Q

What is the significance of the two different conformations of the prion protein?

A

The same sequence produces two diff conformations (normal is alpha-helices whereas disease state contains b-sheets/strands) – therefore it does something completely differently, thus the prion state leads to disease.

165
Q

What do PrP^Sc proteins form?

A

Amyloid fibres

166
Q

What are amyloid fibres?

A

Amyloids are aggregates of proteins that become folded into a shape that allows many copies of that protein to stick together, forming fibrils.

167
Q

What is the clinical significance of amyloid fibres?

A

Pathogenic amyloids form when previously healthy proteins lose their normal physiological functions and form fibrous deposits in plaques around cells which can disrupt the healthy function of tissues and organs.

Such amyloids have been associated with (but not necessarily as the cause of) more than 50 human diseases, known as amyloidoses, and may play a role in some neurodegenerative disorders

168
Q

Fig. 12a

Label the image.

A

PrP^C

169
Q

Fig. 12b

Label the image.

A

PrP^Sc

170
Q

Draw a picture of the PrP^C protein.

A

See Fig. 12a

lots of alpha helices

171
Q

Draw a picture of the PrP^SC protein.

A

See Fig. 12b

lots of beta sheets/strands instead of alpha-helices.

172
Q

What happens if a protein misfolds?

A

If proteins don’t fold properly, they often go through a common state (prions) which causes this disease-like state (beta-sheets) which causes amyloidoses.

173
Q

Give 3 examples of amyloidoses.

A
  • Alzheimer’s disease
  • Type II diabetes (some types)
  • Spongiform encephalopathies
174
Q

What cause Alzheimer’s disease via amyloidoses?

A

Amyloid b-peptide forms abnormal amyloid fibre structure

175
Q

What is the pathophysiology of amyloidoses?

A

Abnormal peptide forms abnormal amyloid fibre structure

176
Q

How do amyloidoses appear on histopathology?

A

Clusters in the cell of abnormally folded proteins

177
Q

Name some clinical syndromes and their fibril subunits of some members of the family of systemic extracellular amyloidoses. (Don’t need to learn but to be aware of)

A

Table 1 - Some members of the family of systemic extracellular amyloidoses

Clinical syndrome: Fibril subunit

  • Primary systemic amyloidosis: Intact immunoglobulin light chains or fragments thereof
  • Secondary systemic amyloidosis: Fragments of serum amyloid A protein
  • Familial Mediterranean fever: Fragments of serum amyloid A protein
  • Familial amyloidotic polyneuorpathy 1: Mutant transthyretin and fragments thereof
  • Senile systemic amyloidosis: Wild-type transthyretin and fragments thereof
  • Familial amyloidotic polyneuropathy II: Fragments of apolipoprotein A-1
  • Haemodialysis-related amyloidosis: B2-Microglobulin
  • Finnish hereditary amyloidosis: Fragments of mutant gelsolin
  • Lysozyme amyloidosis: Full-length mutant lysozyme
  • Insulin-related amyloid: Full-length insulin
  • Fibriniogen a-chain amyloidosis: Fibrinogen a-chain variants
178
Q

Name some clinical syndromes and their fibril subunits of some of the organ-limited extracellular amyloidoses.

A

Table 2 - Some of the organ-limited extracellular amyloidoses

Clinical syndrome: Fibril subunit

  • Alzheimer’s disease: Amyloid b-peptide
  • Spongiform encephalopathies: Full-length prion protein or fragments thereof
  • Hereditary cerebral haemorrhage with amyloidosis: Amyloid b-peptide or cystatin C
  • Type II diabetes: Amylin (islet amyloid polypeptide)
  • Medullary carcinoma of the thyroid: Procalcitonin
  • Atrial amyloidoses: Atrial natriuretic factor
179
Q

Name some diseases and their protein and associated loci of some human brain diseases characterised by progressive misfolding and aggregation of proteins.

A

Table 3 - Some human brain diseases characterised by progressive misfolding and aggregation of proteins

Disease / Protein / Locus

  • Alzheimer’s disease / Amyloid b-protein, Tau / Extracellular plaques, Tangles in neuronal cytoplasm
  • Frontotemporal dementia with parkinsonism / Tau / Tangles in neuronal cytoplasm
  • Parkinson’s disease; dementia with Lewy bodies / a-Synuclein / Neuronal cytoplasm
  • Creutzfeldt-Jakob disease ‘mad cow disease’* / Prion protein (PrP^Sc) / Extracellular plaques; Oligomers, inside and outside neurons
  • Polyglutamine expansion diseases (Huntington’s disease, spinocerebellar ataxias, and so on)* / Long glutamine stretches within certain proteins / Neuronal nuclei and cytoplasm
  • Amyotrophic lateral sclerosis* / Superoxide dismutase / Neuronal cytoplasm

*Figures reproduced from Greenfield’s Neuropathology (copyright Hodder Arnold)

180
Q

Fig. 13

Label the diseases characterised by these histopathologies.

A
  • Alzheimer’s disease
  • Parkinson’s disease; dementia with Lewy bodies
  • Creutzfeldt-Jakob disease ‘mad cow disease’
  • Polyglutamine expansion diseases (Huntington’s disease, spinocerebellar ataxias, and so on)
  • Amyotrophic lateral sclerosis
181
Q

Draw the histopathology for some prion diseases of the brain.

A

See Table 13

  • Alzheimer’s disease / Extracellular plaques, Tangles in neuronal cytoplasm
  • Parkinson’s disease; dementia with Lewy bodies / Neuronal cytoplasm
  • Creutzfeldt-Jakob disease ‘mad cow disease’ / Extracellular plaques; Oligomers, inside and outside neurons
  • Polyglutamine expansion diseases (Huntington’s disease, spinocerebellar ataxias, and so on) / Neuronal nuclei and cytoplasm
  • Amyotrophic lateral sclerosis / Neuronal cytoplasm
182
Q

What are amyloid fibres?

A

Amyloid fibres

  • misfolded, insoluble form of a normally soluble protein
  • highly ordered with a high degree of b-sheet
  • core b-sheet forms before the rest of the protein
  • inter-chain assembly stabilised by hydrophobic interactions between aromatic amino acids
183
Q

What is the molecular detail of amyloidoses?

A

They all adopt a common sort of conformation – form a sort of thing called amyloid fibres

184
Q

What is the relationship between amyloid fibres and the normal proteins?

A

Normal soluble proteins, have misfolded and adopted a completely different conformation which is insoluble. They both have the same amino acid sequence.

185
Q

What is the structure of amyloid fibres?

A

They have this regular repeating pattern – beta strand like structures. Beta sheets can occur normally as part of many proteins, but proteins that are abnormal, tend to adopt this general conformation (not really sure why). These tend to form first before the rest of the protein.

186
Q

What is found between layers pf beta sheets?

A

H-bonds

187
Q

What are beta sheets stabilised by?

A

Their side chains

188
Q

What property of the amino acids involved in the beta-sheets of amyloid fibres allows them to stabilise?

A

They are hydrophobic aromatic residues, which form strong hydrophobic interactions which seem to stabilise them

189
Q

Why is it clinically interesting to know how amyloid fibres are stabilised?

A

Becomes quite important, allows us to think about how we can develop ways to disrupt formation of amyloid fibres – might be a way of treating disease, so ways we can actually interfere with this process and slow down disease progression.

190
Q

Fig. 14 top

Caption this image

A

Amyloid fibres

- misfolded, insoluble form of a normally soluble protein

191
Q

Fig. 14 middle

Caption this image

A
  • highly ordered with a high degree of b-sheet

- core b-sheet forms before the rest of the protein

192
Q

Fig. 14 bottom

Caption this image

A
  • inter-chain assembly stabilised by hydrophobic interactions between aromatic amino acids
193
Q

Draw an image of amyloid fibres.

A

See Fig. 14 top

Amyloid fibres
- misfolded, insoluble form of a normally soluble protein

194
Q

Draw an image of the secondary structure of amyloid fibres.

A

See Fig. 14 middle

  • highly ordered with a high degree of b-sheet
  • core b-sheet forms before the rest of the protein
195
Q

Draw an image of how beta sheets in amyloid fibres are stabilised.

A

See Fig. 14 bottom

  • inter-chain assembly stabilised by hydrophobic interactions between aromatic amino acids

(Sheets on top of each other, side chains hang down or stand up and interact with each other. Aromatic.)

196
Q

What do I need to know?

A

An outline of the forces involved in protein structure and at what level they are important
e.g. H-bonds from backbone peptide bonds in secondary structure

An outline of how proteins fold

Why protein folding is important for function

Why protein misfolding can cause disease
- no need to understand the molecular interactions that stabilise amyloid fibre structure

197
Q

Give an outline of the forces involved in protein structure and at what level they are important

A

See Slides 3-6

198
Q

Give an outline of how proteins fold

NB: Need a really general idea of how proteins fold – not in any detail at all - keyword here is outline

A

See Slides 9-11

199
Q

Explain why protein folding is important for function

A

Structure determines function

200
Q
  • Explain why protein misfolding can cause disease*
  • no need to understand the molecular interactions that stabilise amyloid fibre structure

Rough idea why protein misfolding can cause disease and general awareness of amyloid fibres, so not in molecular detail

A

See Slides 12-14

ESA 1 - Medical Cell Biology & Genetics (21 decks)

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