Amino Acids & Structural Determination Techniques Flashcards
draw the side chains of the polar (hydrophilic) amino acids and give their three letter and single letter codes.
pg 1
draw the side chains of the non-polar (hydrophobic) amino acids and give their three letter and single letter codes.
pg 1
draw the side chains of the acidic amino acids and give their three letter and single letter codes.
pg 1
draw the side chains of the basic amino acids and give their three letter and single letter codes.
pg1
Draw the resonance of a peptide bond. Remember that it has a partial double bond.
pg 2
list three a. a. each that have the greatest preference for:
- Alpha helices
- Beta strands
- Reverse turn
- Alpha helices: Glu, Ala, Leu
- Beta strands: Val, Ile, Tyr
- Reverse turn: Gly, Asn, Pro
Explain how secondary structure can be predicted from the amino acid sequence.
- scan a window of 4-5 residues along a sequence, determining average preference of the amino acids
- assign secondary structural features (alpha helices, beta sheets, reverse turns) based on average preference.
What causes most spontaneous folding of proteins?
the hydrophobic effect.
What method of protein representation is best at showing secondary structures?
Ribbon/ cartoon diagram
What is a benefit of the wire protein diagram?
It is simple
In an alpha carbon trace diagram, what is the distance between the alpha carbons assuming all amino acids are in a trans peptide configuration?
~ 3.8 angstroms
What does the ball and stick protein diagram emphasize?
specific residues.
What does the space filling representation of proteins emphasize?
the surface of a protein.
What does the electrostatic/ charge surface representation emphasize?
The accessible binding surface according to the relative charges on the protein.
XRD (X-ray Crystallography)
What are is the min - max size of a molecule?
Pros?
Cons?
1-100 Angstroms
Pros:
- high resolution/accuracy
- easy for model building
- can result in discovery of multiple states/ configurations of a protein
- solvent interactions are often observed, such as a water shell
Cons:
- Need crystals, can take years
- Difficult diffraction/ phasing
- crystal packing artifacts
- High sample purity and homogeneity
- Requires large sample (milligrams of a protein)
NMR (Nuclear Magnetic Resonance)
What is the min - max size of a molecule?
Pros?
Cons?
1-10 angstroms
Pros:
- Shows great dynamics and kinetic information
- No need to crystalize (faster)
- High resolution
- Accurate and detailed, often including hydrogens.
Cons:
- Expensive as proteins need to be labelled by heavy isotopes
- requires high sample purity and homogeneity
- difficult to apply to computational structural determination
- Not good for membrane proteins
- requires lots of proteins (milligrams)
Cryo- EM (cryogenic electron microscopy)
size range of studied proteins?
Pros?
Cons?
100 to >100,000 angstroms
Pros:
- great for dyamics
- no crystalization
- fast sample preparation
- structure in native state
- purity is less of an issue.
Cons:
- less resolution than XRD or NMR
- very expensive
- May only be visible in their preferred orientations, so the entire structure of the protein may not be viewable.
What are the five steps of XRD workflow?
1) obtain protein (molecular cloning and expression)
2) protein purification (fast protein liquid chromatography)
3) protein crystallization (vapor diffusion)
4) data collection (X-ray diffraction experiment)
5) structure determination (model building)
What is the workflow of cryo - EM?
1) place sample in EM grid
2) pick most viewable particles
3) identify particle alignment and sort them
4) create a low resolution Cryo -EM map
5) build or fit a model
What does a lower resolution area of a protein indicate?
Likely that it is disordered and flexible causing there to be more than one configuration.
what are the psi and phi torsion angles of an optimal peptide bond in a:
- right-handed alpha helix
- left-handed alpha helix
- anti-parallel beta sheet
- parallel beta sheet
Consider Ramachandran plots
- phi (-60) psi (-60)
- phi (60) psi (60)
- phi (-120) psi (90)
- phi (120) psi (150)
Why does glycine have a unique Ramachandran graph compared to other amino acids?
Glycine lacks a side chain so rotating it gives less steric clashes.
Alpha Helices
What function can they have?
What is its signature distinguishing feature?
What is the distance between adjacent amino acids?
Alpha helices can be a molecular spring or a hinge point
They are formed via hydrogen bonding between amino acids within the same strand.
~ 1.5 angstroms
Beta Sheet
what is the distance between adjacent amino acids?
What is its signature distinguishing feature?
3.5 Angstroms
hydrogen bonding between strands.
