Lecture 4: Protein Structure and Folding

Question 1

Describe primary protein structure

Answer 1

the linear sequence of amino acids joined enzymatically in a condensation reaction, forming a backbone consisting of peptide bonds and an alpha carbon at each amino acid. Chain has directionality from N terminus to C terminus

Question 2

Describe secondary protein structure

Answer 2

periodic, regular structures; eg alpha helix, beta strands, turns

Question 3

Describe teritary structure

Answer 3

folding of secondary structures into defined protien motifs and domains

Question 4

Describe quaternary structure

Answer 4

assemble of multiple distinct chains into multi-subunit structures

Question 5

Describe peptide bonds

Answer 5

polar, uncharged bonds that experience resonance, resulting in partial pi-bond character that prevents rotation around the peptide bond causing them to be flat

Question 6

Where in a polypeptide chain is rotation allowed

Answer 6

around the bonds linking the amide and the carbonyl to the alpha carbon, where angles range from -180 to 180, though not all angles are permitted as steric clash is minimized in the trans position

Question 7

Describe the alpha helix structure

Answer 7

A right-handed helix with side chains pointing out where intra-strand hydrogen bonds form between backbone residues down the centre of the helix

carbonyl oxygen from an amino acid forms an H-bond with a NH group 4 residues away (i, i+4)

Question 8

How many residues in a full alpha helix turn

Answer 8

3.6 residues per 360 degree turn; since each residue is 3.6A high, a compact structure is created

Question 9

What determines the properties of an alpha helix

Answer 9

side chains and their non-covalent interactions

Question 10

Describe the beta-strand/sheet structure

Answer 10

H-bonds are formed between the backbone carbonyls and amines of different beta strands while R groups alternate to prevent steric hinderence

• strands can run parallel (will loop over), antiparallel (will not loop), or mixed

Question 11

Describe the beta pleated sheet

Answer 11

side chains point above and below the plane of the sheet

Question 12

Describe a beta turn structure

Answer 12

A 4-residue segment allows a complete 180-degree change in direction that can be found on the surface of globular proteins connecting secondary structures

Question 13

What amino acids are commonly found in beta turns1

Answer 13

Proline most common at position 2; its fixed angle helps initiate a turn

Gly, Asn, and Ser also frequently seen (Asn and Ser due to PTMs and Gly bc its small)

Question 14

What are motifs and domains

Answer 14

commonly seen tertiary structures that often play specific functional roles (eg specific binding motif/ binding domain)

Question 15

What is the zinc finger motif

Answer 15

motif found in DNA-binding proteins that contain zinc ions that coordinate distant side chains to stabilize domains

eg histidine binds to zinc, zinc binds to leucine= bond stabalized

Question 16

What are six commonly seen structural motifs

Answer 16

1) coiled coil
2) helix bindle
3) beta-alpha-beta unit
4) hairpin (reverse turn)
5) Greek key
6) beta-barrel

Question 17

example of a homodimer

Answer 17

alcohol dehydrogenase

Question 18

two examples of heterodimers

Answer 18

hemoglobin, immunoglobulin antibody

Question 19

How does the body tell the difference between different antibody types

Answer 19

different primary sequences dictate different noncovalent interactions

Question 20

Describe x-ray crystallography

Answer 20

some proteins form ordered crystals under varying conditions and these crystals produce diffraction patterns when placed in an X-ray diffractometer that can be interpreted in terms of atomic positions

Question 21

Describe how to read electron density maps

Answer 21

diffraction pattern generates a map, where blank space indicated areas that an X-ray bounced off, aka a nucleus- work backwards and fill in blank space with imagined structure

High resolution obtained by a low distance (low A value)

Question 22

What is the limiting factor in x-ray crystallography

Answer 22

creating crystals

Question 23

Describe nuclear magnetic resonance (NMR)

Answer 23

-carried out on proteins in solution
- based on the nuclear spin of 1H, 13C, or 15N nuclei that can be measured in a strong, static magnetic field
- the absorption of radiation can be used to deduce the environment of a nucleus to determine a proteins structure
- monitors conformational changes, including folding and interactions with other molecules

Question 24

Describe the 1D vs 2D NMR spectra of a protein

Answer 24

1D represented on a graph or intensity vs chemical shift and peaks are observed

2D, a dot is the intersection of 2 peaks representing different amines or amines. Red dots represent changes observed upon binding (conformational change)

Question 25

What is the function of cryo-electron microscopy

Answer 25

Allows larger complexes to be visualized (over 100kDa) and a 3D structure of a single protein to be built by averaging out multiple structures

Question 26

Describe the process of cryo-electron microscopy

Answer 26

A thin layer of protein solution is prepared on a fine grid and flash-frozen to trap molecules in different orientations High-powered microscopes are used to measure a beam of electrons that pass through the protein sample; diffraction of the beam of electrons can be used to determinethe structure

Question 27

Describe AlphaFold and AlphaFold3

Answer 27

AI model able to predict protein structures and molecular interactions that was developed as a free server by Google DeepMins and Isomorphic Labs Limited size and processing power

Question 28

What are chromaphores and what are some important ones found in proteins?

Answer 28

molecules that contain conjugated double bonds that absorb UV light at specific wavelengths. Aromatic rings and amide carbonyls are important and found in proteins

Question 29

How can chromaphores be used to help determine protein structure

Answer 29

the structure of a protein influences the accessibility of chromophores to light absorption

Question 30

What are fluorophores

Answer 30

molecules that absorb and emit light; used for cellular localization

Question 31

What allows for tryptophan fluiorescence

Answer 31

The presence of an infole ring allows Trp to absorb when excited with UV light (270-295nm) and then emit light between 310-355nm

Question 32

What determines the exact wavelength of light emitted by tryptophan

Answer 32

the polarity of its environment in a protein in a polar environment, the fluorescence is 'red-shifted', wavelengths are longer and less intense (lower energy)

Question 33

How can pH affect tryptophan fluorescence

Answer 33

pH can lead to changes in structure which will be reflected as changes in the level or fluorescence

Question 34

How can the location of tryptophan in a protein be determined based on the wavelength of fluorescence it emits

Answer 34

a polar, surface protien will be 'red shifted', higher wavelength less energy, around 350nm a nonpolar, buried protein will be 'blue-shifted', lower wavelength, higher energy, around 310nm

Question 35

What are chaperone proteins

Answer 35

proteins that assist in folding by preventign the aggregation of newly synthesized and unfolded proteins by binding to exposed hydrophobic regions

Question 36

What is protein denaturation

Answer 36

the disruption of the weak forces that hold together protein structure by external stressed that results in the loss of structure and function

Question 37

Name six diseases resulting from misfolded proteins

Answer 37

- Altzheimers disease - Familial amyloidotic polyneuropathy - cancer - mad cow disease - hereditary emphysema - cystic fibrosis

Question 38

What is the difference between the wildtype and scrapie prion protein and what triggers this change

Answer 38

wildtype has alpha helices, scrapie has beta sheets though both have same starting structure trigger is some unknown environmental change