Cell and Molecular - Lecture One

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Name three diseases caused by protein misfolding
Alzheimer's, Parkinson's, Cancer
1 of 50
Describe an alpha helix conformation
Spiral conformation in which every backbone NH2 group donates a hydrogen bond to the C=O group of an amino acid 3 to 4 residues earlier along the protein
2 of 50
How many nm and residues does a full turn of an alpha helix span?
0.5nm and 3.6 residues
3 of 50
Which amino acids have high alpha helix forming propensities?
Methionine, alanine, leucine, glutamate and lysine
4 of 50
Which amino acid have low alpha helix forming propensities?
Proline and glycine
5 of 50
Describe a beta-sheet conformation
A secondary structure formed by hydrogen bonds between adjacent beta-strands
6 of 50
True or false - aromatic/hydrophobic residues are found in the middle of beta-sheets?
True
7 of 50
Name some amino acids that are likely to be found in the middle of beta-sheets
tyrosine, phenylalanine, tryptophan, valine, isoleucine
8 of 50
What is the role of the enzyme zymase?
It catalyses the conversion of sugar into ethanol and CO2
9 of 50
Describe the structure of the Src tyrosine kinase
It's multidomain - it has a large kinase domain, a small kinase domain, and 2 regulatory domains - SH2 and SH3
10 of 50
True or False - the small kinase domain is made up of mostly alpha helices?
FALSE - the small kinase domain is made up of mostly Beta-strands, and the large kinase domain is made up of mostly alpha helices.
11 of 50
What are the three interactions between proteins?
1. Ionic interactions - between +ve and -ve ions
12 of 50
How do peptides fold into a protein?
They fold so the hydrophobic amino acids are hidden inside the structure to form a hydrophobic core region, where as the polar side chains are found on the outside as they make hydrogen bonds with surrounding water molecules
13 of 50
What is the primary structure?
The linear sequence of amino acids
14 of 50
What is the secondary structure?
How the amino acids fold to give a particular shape e.g. alpha helix, beta-sheet or random coil
15 of 50
What is the tertiary structure?
They way secondary structures pack together within a protein
16 of 50
What is the quaternary structure?
The relationship between individual proteins into a multimeric complex
17 of 50
How is the primary structure mainly inferred?
From the DNA sequence
18 of 50
Name another way primary structure can be inferred?
By sequencing the amino acids
19 of 50
What is the role of Edman degradation?
Used to work out the secondary structure of a protein by amino acid sequencing
20 of 50
What is the role of mass spectrometry?
Used to work out the primary structure of a protein by amino acid sequencing
21 of 50
Explain how Edman degradation takes place
It's a chemical disruption - PITC binds and disrupts the terminal amino acid, you then run HPLC to work out the MW of the amino acid.
22 of 50
What is the main way in which secondary structure is predicted?
Use the primary sequence to predict secondary sequence - this can be done by de novo or by alignment with proteins of known structures
23 of 50
Describe the structure of a coiled-coil?
Two amphipathic alpha helices wrap around each other so the hydrophobic sides come together in the centre and polar faces are exposed outways
24 of 50
Where would you typically find coiled coils?
In elongated aggregated proteins
25 of 50
Describe the structure of insulin
3 alpha helical elements and random coils
26 of 50
What is the role of Circular Dichroism?
To estimate secondary structure
27 of 50
What range on the UV spectrum is CD done in?
The far UV spectrum at 190-250nm
28 of 50
On the CD spectrum, how many peaks would be characteristic of an alpha helix?
2
29 of 50
On the CD spectrum, how many peaks would be characteristic of a beta-sheet?
1
30 of 50
What is the definition of CD?
A differential absorption of circularly polarised light
31 of 50
True or false - you can look at how various conditions affect protein structure e.g. pH and temp using CD?
True
32 of 50
How does CD tell us about the stability of a protein?
Protein unfolding or melting is followed by CD at different temperatures, it causes the protein to uncoil, the less stable a protein the faster it loses its CD characteristics upon heating
33 of 50
Describe how x-ray crystallography tells us about the structure of a protein?
A high energy focused beam of x-rays is fired through a protein crystal - look at the diffraction pattern to determine structural elements
34 of 50
True or False - NMR uses isotopes such as C13?
True - the isotopes have different numbers of protons and neutrons and hence the nucleus has a slight wobble in the spin
35 of 50
What is chemical shift?
Chemical shift is when different protons resonate at different frequencies depending on the local environment
36 of 50
How would you introduce the isotopes into the structure of a protein?
You would produce them recombinantly in bacteria where the whole nutrient source is the isotopes, so all proteins are labelled
37 of 50
True or false - NMR is a repetitive process?
T - you usually arrive at several possible structures represented as an ensemble
38 of 50
What is TEM?
Transmission Electron Microscopy - protein is placed in heavy metal that electrons cannot penetrate, this is used to build up an overall structure
39 of 50
Give an example of a heavy metal?
Uranyl acetate
40 of 50
What is cryo-EM?
Proteins are frozen in vitreous ice, by shinning electrons onto the frozen structure you can determine the shape of the protein
41 of 50
True or false - it is easier to process more radial symmetrical structures?
True - an example is a virus.
42 of 50
Give three advantages of CD?
Its cheap, quick and theres no size limit
43 of 50
Give two disadvantages of CD?
Limited dynamics and very low resolution (secondary structure only)
44 of 50
Give three disadvantages of crystallography?
It's very costly, its slow and dynamics can be limited due to crystallisation artefacts
45 of 50
Give two advantages of crystallography?
No size limit (although it gets harder as you get bigger) and it has the highest resolution overall - 1A
46 of 50
Give four disadvantages of NMR?
Very costly, analysis is slow, isotopic label size can be a problem, size limit of 50kDa
47 of 50
Give two advantages of NMR
Good dynamics, reasonable resolution - 2A
48 of 50
True or False - EM is quite cheap?
False - it's quite costly
49 of 50
Does EM have a size limit?
No
50 of 50

Other cards in this set

Card 2

Front

Describe an alpha helix conformation

Back

Spiral conformation in which every backbone NH2 group donates a hydrogen bond to the C=O group of an amino acid 3 to 4 residues earlier along the protein

Card 3

Front

How many nm and residues does a full turn of an alpha helix span?

Back

Preview of the front of card 3

Card 4

Front

Which amino acids have high alpha helix forming propensities?

Back

Preview of the front of card 4

Card 5

Front

Which amino acid have low alpha helix forming propensities?

Back

Preview of the front of card 5
View more cards

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