Protein Life Cycle

Question 1

Cytoplasmic Crowding

Answer 1

Two fold effect: retards unfolding but enhances aggregation

Hydrophobic residues can be pushed aside and form aggregates instead of actual protein

Question 2

Physical factors that influence unfolding equilibrium

Answer 2

Denaturing: pH (lysosome, acidosis, bone remodeling, nerve synapses)
Ionic strength: salt
Pressure: Proteins running through blood
Temperature
Osmotic pressure
Urea (kidneys)

Question 3

Osmolytes

Answer 3

Cause nonspecific effect on all proteins they encounter

Higher concentrations

Enhance protein folding
AA: Proline, Alanine, Taurine
Polyols: Sorbitol, Glycerol
Methylamines: TMAO, betaine

Question 4

Kidney Cells and Stress

Answer 4

Cell up/downregulates osmolytes to compensate for changing urea and salt concentrations

Question 5

ER and Protein Processing

Answer 5

Oxidizing: Disulfide bonds and glycosylation

Increase protein stability

Question 6

Covalent Osmolytes

Answer 6

Very effective for protein stabilization

~50% of proteins are naturally glycosylated

Question 7

Allergens

Answer 7

Proteins that are so stable they can survive digestion.
Must be >25 AA for it to be allergen

Ovomucoid: egg white allergen. (disulfide bonds, glycosylation)

You cannot unfold these proteins, meaning there’s a recognizable chunk that your immune system cant recognize.

Things larger than 25 AA, the immune system can recognize, which is what causes the allergic reaction.

Question 8

Chaperones

Answer 8

Protein that the cell makes to help other proteins fold up

Interact often with misfolded or unfolded

Heat shock proteins

Can transport across membrane
Control activities of some proteins

Question 9

Chaperonin

Answer 9

Class of chaperones that assist with folding of proteins using ATP

Central cavity
TriC = structure of humans
GroEL = E. coli

Interacts with about 10% of new proteins

Question 10

Protein Turnover

Answer 10

1-2% degradation each day

Question 11

Proteasome

Answer 11

Short half life, transient function, abnormal or damaged

Question 12

Lysosome

Answer 12

Longer half life, “housekeeping” function, membrane

Question 13

Lysosomal degradation

Answer 13

Autophagy: nonselective, slow, constitutive

Chaperone-mediated autophagy: selective, 30% of cytosolic proteins, hsc73 binding to KFERQ motifs (bind to, recognize, haul to lysosome), unfolded protein transported into lysosome

Question 14

Cathepsin Proteases

Answer 14

> 12, found in the lysosome
Constitutive and tissue specific (some are protein specific, some found in all lysosomes)
broad, overlapping substrate specificities

Question 15

Proteasome

Answer 15

Cytosol and nucleus
Neutral pH
Ubiquitin tags on protein targets
Requires ATP

Question 16

Proteosome Pathway

Answer 16

Ubiquitin binds to protein through ATP use
Ubiquitin-protein is bound to proteasome
ATP used again to degrade protein
Ubiquitin is recycled

Question 17

Ubiquitin Tagging

Answer 17

E1 + ubiquitin: 1 activator protein, ATP required
E2 + ubiquitin: Several carrier isoforms
E3 + target protein + ubiquitin

E3 special because recognizes target protein and ubiquitin
>500 ligase proteins

Question 18

Ubiquitinylation

Answer 18

Covalent attachment at lysine side chain (N)
Amide bond with Ubq (C)

1 Ubq is not enough: need 4 in order to be broken down by proteosome

Question 19

Structure of Proteosome

Answer 19

Stacked rings
Three separate protease in middle rings
Each recognizes a different AA cleavage site

Question 20

Protein Turnover (AA)

Answer 20

75% of free amino acids are recycled into new proteins
25% AA are metabolized
> 1. Nitrogen excreted as urea
>2. Carbon components “burned” for energy

Question 21

Proteasome and Cancer

Answer 21

Induces death of cancer cells
Few effects on normal cells

Bortezomib: approved for relapsed multiple myeloma
Inhibits activity of proteasome

Question 22

ER: Unfolded Protein Response

Answer 22

Types of stress: 
unfolded proteins in ER (mutation or improper glycosylation)
ROS or UV damage
Low amino acids
Heat (fever)
Hypoxia
High free fatty acids

Stress sensed, cell physiology changes >
recovery or apoptosis

Question 23

BiP

Answer 23

Chaperone that senses stress
Normally binds sensor proteins
Inactive: BiP is bound to sensor
Active: BiP is unbound, can interact and send signals

At lease three sensor proteins, each with its own pathway of sending messages to the nucleus.

Sensor stretches across ER membrane & is released in ER lumen to interact with unfolded proteins

Question 24

Cellular Responses

Answer 24

1. Inhibit global translation (quit making more protein, only going to make the problem worse)
2. Activate transcription/translation of:
   ERAD proteins: that assist with degradation of misfolded proteins in ER (includes ubiquitin system)
   Chaperones (upregulates)
   AA transporters (upregulates- to have more building blocks to build proteins from)
   Oxidative stress protection

Question 25

Ex. of temperature causing protein denaturation

Answer 25

Tachycardia develops when a child runs a fever

Question 26

Ligand

Answer 26

Includes co-factors
Specific to only some proteins, will not bind to all

Binding a ligand will stabilize the protein conformation

Question 27

Chemical environment of body

Answer 27

pH and ionic strength frequently change

Some areas are designed to unfold proteins (intestinal tract, lysosome)

The body is influenced by a high density of macromolecules

Question 28

Aquaporins

Answer 28

Transport water

Question 29

Catalase and superoxide dismutase

Answer 29

Regulate redox reactions

Question 30

Multi-drug transporters

Answer 30

Transport hydrophobic ligands across membrane

Question 31

Histidine (sensing)

Answer 31

charged side chain is able to sense pH changes near 7

Question 32

Hemoglobin

Answer 32

Cannot bind O2 without heme

Can bind up to 4 O2 molecules