Module 3 - hierarchy of protein structure and folding

Question 1

Why can proteins change shape so easily?

Answer 1

they are held together by weak interactions

Question 2

What are the 4 hierarchial levels of protein structure?

Answer 2

primary - amino acid sequence
secondary - a helices, b strands, or b turns
tertiary - arrangement of all atoms in the polypeptide chain
quaternary - arrangement of subunits

Question 3

How are alpha helices stabilized?

Answer 3

intrastrand hydrogen bonding between the carbonyl oxygen of one amino acid and the hydrogen of the amide four residues away

Question 4

How do the amino acid dipoles affect the dipole of the overall alpha helice?

Answer 4

they all add together in the same direction to create a net dipole with partial charges on both ends (positive at the N terminus, negative at the C terminus)

Question 5

Describe amphipathic alpha helices

Answer 5

amino acids with hydrophilic or hydrophobic properties are positioned every 3-4 residues so when the helix forms they are one their own respective faces

• located on the outside of globular proteins, with a hydrophobic face facing into the protein (ex: apolipoprotein)
• also located as transmembrane proteins, with the hydrophobic face pointing out toward the membrane and hydrophilic residues pointing in (rhodopsin)
• helical wheels can be used to predict the presence of amphipathic alpha-helices based on the spacing of hydrophilic and hydrophobic amino acids

Question 6

How do parallel and antiparallel beta sheets form? Which is more stable?

Answer 6

parallel: when peptides are stacked on top of each other, the carboxyl groups and amino groups line up vertically, so H bonds form diagonally between them

antiparallel: when peptides are stacked on top of each other, the carboxyl groups are directly in line with the amino groups, so H bonds form vertically

antiparallel are more stable than parallel, and they are also more common in proteins (called B pleated sheets)

Question 7

What is a beta turn vs a loop?

Answer 7

turn: small 3-4 amino acid turns
loop: on protein surfaces, more amino acids in length

Question 8

Where are the general areas for alpha helices, beta turns, parallel beta sheets, and antiparallel beta sheets on a Ramachandran plot?

Answer 8

alpha helices around the middle, beta turns at the top center, both beta-sheets at the top left but antiparallel are slightly higher and to the left

Question 9

How is a Ramachandran plot developed?

Answer 9

from measuring phi and psi angles of existing structures

Question 10

What is the difference between tertiary structures that are a/B and a+B?

Answer 10

a/B - both mixed together in the same domain
a+B - different domains (half protein is a, half is B)

Question 11

What domains are present in the structure of pyruvate kinase?

Answer 11

all 4 types - short helical, a/B, B, and a+B

Question 12

What are the 4 special types of tertiary structures?

Answer 12

four-helix bundle, greek key fold, Rossmann fold, and TIM Barrel fold

Question 13

What is the four helix bundle?

Answer 13

normal “squiggly line” formation with alpha helices on the straight lines

Question 14

What is the greek key fold?

Answer 14

beta strands that resemble the greek key

up, left, down, left, up, way right, down

Question 15

What is the Rossmann fold?

Answer 15

alternating alpha helices and beta strands, two regions

up, right, down, right, up, right, down, right, up
WAY left, down, left, up, left, down, left, up, left, down, left, up

Question 16

What is the TIM Barrel fold?

Answer 16

alternating alpha helix/beta strand fold
just a squiggly line, but alternating a/B

Question 17

How can tertiary structures be stabilized?

Answer 17

disulfide bonds between cysteines or by metal ions like zinc fingers

Question 18

Describe the quaternary structure of keratin

Answer 18

homodimer of two helical polypeptides that form a coiled-coil held together by disulfide bridges

Question 19

Describe the quaternary structure of collagen

Answer 19

three helical subunits held together by cross-links of hydroxylated lysines

Question 20

Describe the quaternary structure of G protein

Answer 20

heterotrimeric quaternary complex with three distinct polypeptides (a1b1g1)

Question 21

Describe the quaternary structure of hemoglobin

Answer 21

Heterotetrameric complex containing two copies each of an a and b subunit, giving rise to an a2b2 heterotetramer. Each subunit contains an oxygen-binding heme.

Question 22

Describe the quaternary structure of immunoglobulins

Answer 22

aka antibodies, two copies each of related protein subunits (two heavy chains and two light chains) with antigen binding sites at the end of the light chains

the variable and constant domains are all-b structures called Ig folds

Question 23

Protein unfolding is a —– ——-.

Answer 23

cooperative process

Question 24

Describe the Anfinsen experiments and what was learned from them.

Answer 24

• urea and BME were added to fully unfold a protein
• when both were removed at the same time, the protein refolded with the correct disulfide bonds
• when BME was removed first, it refolded incorrectly.

Why?
- BME reduces sulfurs and breaks disulfide bonds (tertiary structure)
- urea denatures hydrogen bonds (secondary structure)
- removing BME first causes wrong disulfide bonds to form because the correct hydrogen bonds were not in place first

What was learned?
- primary amino acid sequences contain all structural information required for protein function using ribonuclease A (RNaseA)

Question 25

Before alpha helices form completely, they are partially formed early on as part of structures called…

Answer 25

molten globules

Question 26

What is the hydrophobic collapse model?

Answer 26

suggests that the clustering of hydrophobic side chains drives the formation of a hydrophobic core which facilitates secondary and tertiary structure formation

Question 27

What does the folding funnel visually illustrate?

Answer 27

there is not a single starting point or single path taken to achieve the final folded state of a protein

Question 28

What are the two examples of chaperone proteins?

Answer 28

GroEL/GroES protein complex and the Hsp70 (heat shock protein 70)

Question 29

How does the GroEL/GroES protein complex function?

Answer 29

ATP opens top and unfolded protein enters, closes and folds, conformational change releases a protein that was previously folded in the bottom and ATP causes and new unfolded protein to enter the bottom, conformational change opens the top and the folded protein is released, etc.

Question 30

How does the Hsp70 chaperone protein work?

Answer 30

• when bound to ATP, the Hsp70 undergoes a conformational change to reveal hydrophobic groups that bind to an unfolded protein
• ATP is hydrolyzed and Hsp70 closes on the protein to help it fold
• ADP is released, and Hsp70 opens again to release the partially refolded protein (can be repeated until folding is complete)

Question 31

Some diseases are caused by proteins that misfold (but have no amino acid changes). What are some examples, and the differences between them?

Answer 31

Cystic fibrosis - misfolded proteins degrade and cause loss of function
Huntington’s disease - misfolded proteins aggregate and cause a gain of function
Creutzfeld-Jakob disease - a different protein conformation aggregates and causes gain of function