Amino Acids & Structural Determination Techniques

Question 1

draw the side chains of the polar (hydrophilic) amino acids and give their three letter and single letter codes.

Answer 1

pg 1

Question 2

draw the side chains of the non-polar (hydrophobic) amino acids and give their three letter and single letter codes.

Answer 2

pg 1

Question 3

draw the side chains of the acidic amino acids and give their three letter and single letter codes.

Answer 3

pg 1

Question 4

draw the side chains of the basic amino acids and give their three letter and single letter codes.

Answer 4

pg1

Question 5

Draw the resonance of a peptide bond. Remember that it has a partial double bond.

Answer 5

pg 2

Question 6

list three a. a. each that have the greatest preference for:
- Alpha helices
- Beta strands
- Reverse turn

Answer 6

• Alpha helices: Glu, Ala, Leu
• Beta strands: Val, Ile, Tyr
• Reverse turn: Gly, Asn, Pro

Question 7

Explain how secondary structure can be predicted from the amino acid sequence.

Answer 7

• 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.

Question 8

What causes most spontaneous folding of proteins?

Answer 8

the hydrophobic effect.

Question 9

What method of protein representation is best at showing secondary structures?

Answer 9

Ribbon/ cartoon diagram

Question 10

What is a benefit of the wire protein diagram?

Answer 10

It is simple

Question 11

In an alpha carbon trace diagram, what is the distance between the alpha carbons assuming all amino acids are in a trans peptide configuration?

Answer 11

~ 3.8 angstroms

Question 12

What does the ball and stick protein diagram emphasize?

Answer 12

specific residues.

Question 13

What does the space filling representation of proteins emphasize?

Answer 13

the surface of a protein.

Question 14

What does the electrostatic/ charge surface representation emphasize?

Answer 14

The accessible binding surface according to the relative charges on the protein.

Question 15

XRD (X-ray Crystallography)
What are is the min - max size of a molecule?
Pros?
Cons?

Answer 15

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)

Question 16

NMR (Nuclear Magnetic Resonance)
What is the min - max size of a molecule?
Pros?
Cons?

Answer 16

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)

Question 17

Cryo- EM (cryogenic electron microscopy)
size range of studied proteins?
Pros?
Cons?

Answer 17

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.

Question 18

What are the five steps of XRD workflow?

Answer 18

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)

Question 19

What is the workflow of cryo - EM?

Answer 19

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

Question 20

What does a lower resolution area of a protein indicate?

Answer 20

Likely that it is disordered and flexible causing there to be more than one configuration.

Question 21

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

Answer 21

Consider Ramachandran plots

• phi (-60) psi (-60)
• phi (60) psi (60)
• phi (-120) psi (90)
• phi (120) psi (150)

Question 22

Why does glycine have a unique Ramachandran graph compared to other amino acids?

Answer 22

Glycine lacks a side chain so rotating it gives less steric clashes.

Question 23

Alpha Helices
What function can they have?
What is its signature distinguishing feature?
What is the distance between adjacent amino acids?

Answer 23

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

Question 24

Beta Sheet
what is the distance between adjacent amino acids?
What is its signature distinguishing feature?

Answer 24

3.5 Angstroms
hydrogen bonding between strands.