Lecture 18 Sjogren

Question 1

Molecular Chaperones

Answer 1

assist the covalent folding or unfolding of proteins

assembly or disassembly of macromolecular structures (oligomerization or aggregates)

Question 2

Examples of Chaperones

Answer 2

HSP60 - GroEL/GroES

HSP70 - DnaK

HSP90 - HtpG

Question 3

Disorders in Chaperones

Answer 3

affect muscle, bone, and CNS

Question 4

Problematic Chaperones

Answer 4

partially folded or misfolded proteins are bad –> aggregation

• exposure of the hydrophobic residues and unstructured polypeptide backbone
• aggregation results in amorphous structures

Question 5

Overview of Proteasome Complexity through PTM

Answer 5

Genome (20-25,000 genes) –> Alternative promoters and alternative splicing –> Transcriptome (100,000 transcripts) –> PTM –> Proteome (1,000,000 proteins)

Question 6

PTM Functions

Answer 6

Proteolytic Processing & Conformational Change –> Activation

PTM-dependent proteolysis –> Degradation

PTM-dependent recognition –> Activation, interaction, localization, and secretion

Reversible-site PTM –> Dynamic regulation or modulation

Question 7

Purpose of Protein Degradation

Answer 7

misfolded proteins

dietary proteins to supplement amino acids

regulation of cellular processses

Question 8

Protein Degradation Overview

Answer 8

Controls:
- blood clotting
- processing pro forms of proteins
- cell matrix proteolysis
- replication and transcription

Eliminates:
- misfolded or unfolded proteins
- damage proteins
- large aggregates (cause Alzeheimer, Parkinson, Huntington, ALS)

Question 9

Protein Degradation Processes

Answer 9

Ubiquitin Proteolysis
- proteins that are targeted for degradation

ERAD
- misfolded protein in ER is translocated to cytoplasm by chaperones entering the ubiquitin process

Lysosomal Degradation
- membrane bound organelles containing proteases that will degrade exogenous proteins

Autophagy
- maintains normal functioning by protein degradation (appears in stress)

Apoptosis
- programmed cell death

Question 10

Autophagy and Lysosomal Degradation

Answer 10

• uptake and recycling of nutrients and receptors
• merges with autophagic pathway
• can degrade exogenous or endogenous proteins
• double membrane structure forms by vesicle nucleation around cytoplasmic contents and forms a autophagosome
• fusion of the autophagosome with lysosome becomes a autolysosome
• degradation produces amino acids, fatty acids that can be recycled

Question 11

Defective Autophagy

Answer 11

• When the accumulation of autophagosomes becomes larger than the autophagic degradation, this can lead to neurodegeneration and alzheimer’s

Causes of Increase autophagosomes
- aberrant activation of autophagy
- disruption of autophagosome-lysosome function
- inhibition of lysosome acidification

Question 12

Ubiquitiin-Proteasomal System

Answer 12

• cell control, cell differentiation, and stress response
• must be covalently modified on a lysine

3 Steps
- E1 (activating): ATP hydrolysis to add ubiquitin to a cysteine
- E2 (conjugating): receives ubiquitin on cysteine (transfers from E1 to E2)
- E3 (ligase): specific recognition of protein to be degraded as it transfers the ubiquitin from E2 to the lysine of the substrate

MUST CONTAIN AT LEAST 4 UBIQUITIN

Question 13

Proteasome

Answer 13

20S + 19S = 26S Proteasome

Question 14

20S

Answer 14

core complex consisting of alpha and beta subunits

able to degrade short unfolded non-ubiquinated proteins

trypsin, caspase, and chymotrypsin activity

Question 15

19 S

Answer 15

contains 2 molecules

regulatory cap consisting of multiple subunits

responsible for deubiquination, protein unfolding, and feeding the proteasome

Question 16

Oxidative Stress to Proteasome

Answer 16

ECM29 sequests the 19S caps to be bound and held to the HSP70 protein

20S is free to degrade oxidized proteins

Question 17

Key Difference between Lysosome and UPS

Answer 17

Target Specificity

Degron: recognition sequence or structure for an E3 ligase

Ex.
- N end rule: N terminal sequence of protein is recognized and the 2nd residue can be destabilized (Arg, Leu, Phe)

• PEST Sequences (Proline, Glutamic Acid, Serine, Threonine
• PTM through phosphorylation

Question 18

Protein Misfolding in Cancer

Answer 18

• unregulated cell division requires increased protein synthesis
• cancer cells are susceptible to drugs that alter proteostasis

MORE SUSPECITBLE TO PROTEOLYSIS THAN NORMAL CELLS