Protein Structure Flashcards
The misfolding of what protein causes Alzheimers?
Amyloid B-peptide or Tau in the ER
The misfolding of which protein causes Parkinsons?
a-synuclein in the cytosol
The misfolding of which protein causes Cancer?
p53 in the cytosol
How many different amino acids are there?
20
What are the two major protein structures?
Alpha helix
Beta sheet
How can the alpha helix be represented?
Backbone
Sticks
Ribbons
Space filling
Where was the alpha helix first seen?
In the protein alpha keratin
Where do hydrogen bonds form in an alpha helix?
Between every forth peptide bond
So on C=O is linked to a N-H
How often do alpha helixes have a complementary turn?
Every 3.6 amino acids
Where are alpha helixes abundant in the cell?
Cell membranes ie in transport proteins or receptors
Which amino acids do not favour helix forming?
Proline and Glycine
Beta Sheets can run in what two directions?
Parallel or anti-parallel
Which amino acids are usually found in the middle of the B sheets and why?
Aromatic residiues - Tyr, Phe, Trp, Val and Ile
They are hydrophobic
What is always at the start of a protein?
The N terminus - NH2
COOH is always at the end
What are the four subunits of Src tyrosine kinase?
SH3 domain
SH4 domain
Small kinase domain
Large kinase domain
In general, more complex organisms’ proteins have more what?
They have additional domains
What are the 3 types of non covalent weak bonds that can hold proteins together?
Hydrogen bonds
Electrostatic attraction
Van der Waals Attractions
Where do disulphide bonds form?
Inbetween two adjacent cysteine residues
Where do polar side chains tend to form in a folded protein?
On the outside of the folded protein so they can interact with water
The core of a folded protein can be known as what?
Hydrophobic
How can primary sequence be determined?
Inferred from DNA sequence
Determined directly by amino acid sequencing using Edman degredation (chemical degredation)
Mass Spectrometry (physical degredation)
What chemical does edman degredation use?
Phenylisothyocyanate (PITC)
Adding PITC to an amino acid chain forms what?
Phenylthiocarbamoyl derivative with the N terminus
What is formed when the N terminus is cleaved?
Cyclic compound of phenylthiohydantoin forms
PTH amino acid
This does not change the amino acid
Why is Edman degredation useful?
The protein is not damaged
How is Edman degredation carried out?
In cycles ie each cycle for each amino acid
What are the techniques to obtain real protein structure?
Circular dichroism
Xray crystallography
NMR
Electron Microscopy
Circular Dichroism is used for what?
Estimating the secondary structure
How does CD work?
CD if the differential absorption of circulatory polarised light of pure protein structure
CD spec in the spectral region (190 - 250nm) reveals secondary structure
Different structure give different characteristic shapes of the CD spectrum
It gives a percentage for each secondary structure but no information on the arrangement
How can CD tell us about the stability of the protein?
The less stable it is faster it will lose its CD characteristics upon heating
How can CD tell us if the protein can reform after being heated?
After cooling the CD should show the original structure
What is calmodulin?
A small calcium binding protein which activates protein kinases and other proteins
What is NMR useful for?
Understanding the dynamics of protein structures
ie how proteins change upon binding
How does xray crytsallography work?
Proteins are first crystallised
High energy beam of Xrays is fired through the crystal
Some are deflected giving rise to diffraction patterns
How much can set up of crystallography cost?
In house = £200,000
High resolution, synchrotonous based crystallography = £300,000,000
Why is transmission electron microscopy useful?
Looking at large protein structures
Looking at actin filaments
What does EM use?
A negative stain eg heavy metal for shading
What is cryo-electron microscopy?
Uses liquid nitrogen to freeze the specimen
No shading is used just differences in electron density
The more ordered and symetrical the structure, the easier the averaging process
eg viruses have radial symetry
