Functions and Dysfunctions of Protein Processing

Question 1

Types of Mutations (4)

Answer 1

1. Silent: does not change the amino acid.
2. Missense: changes amino acid within the protein.
3. Nonsense: changes codon into stop codon.
4. Frameshift: changes the amino acid sequence.

Question 2

Duchenne Muscular Dystrophy

Answer 2

Frameshift mutation leading to dystrophin gene, resulting in little/no expression of dystrophin protein.
OOF mutations lead to severe DMD. In frame deletions result in more mild forms.
Leads to muscle wasting. Patients usually die from respiratory failure.

Question 3

Eukaryotic mRNA contains (3)

Answer 3

1. Codons.
2. 7-methylguanosine cap at 5’ end.
3. Poly A tail at 3’ end.

Question 4

Aminoacyl tRNAs

Answer 4

Complex of tRNA with amino acid. AA is esterified at the 3’-end of cognate tRNA.

Question 5

Aminoacyl tRNA synthetases

Answer 5

Catalyzes activation of AA with tRNA.

Question 6

Ribosomal complex sites

Answer 6

A: acceptor site. mRNA accepts the tRNA.
P: peptidyl site. Aminoacyl tRNA is attached.
E: empty site. Occupied by empty tRNA before exiting ribosome.

Question 7

Sickle-Cell Anemia

Answer 7

Missense mutation that substitutes Val for Glu. This alters conformation of HbA that deforms the RBCs. These RBCs have poor oxygen capacity and will clog capillaries.

Question 8

Translation steps (3)

Answer 8

Occurs in 5’ —> 3’ direction.

1. Initiation: formation of mRNA, small ribosomal subunit and initiator tRNA.
2. Elongation: activated AA attached to initiating Met via peptide bod.
3. Termination: peptide chain is released from ribosome.

Question 9

Polysomes

Answer 9

Clusters of ribosomes that simultaneously translate a single mRNA, each synthesizing a polypeptide.

Question 10

Prokaryotic Elongation Inhibitors (5)

Answer 10

1. Tetracycline
2. Chloramphenicol
3. Clindamycin
4. Erythromycin
5. Streptomycin

Question 11

Tetracycline

Answer 11

Binds to the small subunit blocking entry of aminoacyl-tRNA to ribsomal complex.

Question 12

Chloramphenicol

Answer 12

Inhibits peptidyl transferase.

Question 13

Clindamycin and Erythromycin

Answer 13

Binds to large subunit which blocks the translocation of the ribosome.

Question 14

Streptomycin

Answer 14

Binds to small subunit and interferes with the binding of fmet-tRNA. Interferes with the small and large unit association.

Question 15

Eukaryotic Elongation Inhibitors (4)

Answer 15

1. Cycloheximide
2. Diphtheria toxin
3. Shiga toxin
4. Ricin

Question 16

Cyclohexamide

Answer 16

Inhibits peptidyl transferase.

Question 17

Diphtheria toxin

Answer 17

Inactivates GTP-bound eEF-2, interfering with ribosomal translocation.

Question 18

Shiga toxin and Ricin

Answer 18

Binds to large subuit, blocking the entry of aminoacyl-tRNA to ribosomal complex.

Question 19

Euk/Prok Elongation Inhibitor

Answer 19

Puromycin. Causes premature chain termination. Resembles the 3’ end of aminoacylated-tRNA. Enters the A site and is added to the growing chain, casing a premature chain release.

Question 20

Cytoplasmic pathway for protein sorting

Answer 20

For proteins destined for cytocol, mitochondria, nucleus and peroxisomes.
Protein synthesis is solely done on a free ribosome in the cytoplasm.

Question 21

Secretory pathway for protein sorting

Answer 21

For proteins destined for ER, lysosomes, plasma membranes or secretion.
Translation begins on free ribosomes but terminates on ribosomes sent to ER. First 20 AAs of of the polypeptide have ER targeting signal sequences.

Question 22

TOM and TIM

Answer 22

TOM - transporter outer membrane of mito.

TIM - transporter inner membrane of mito.

Question 23

Heat shock proteins 70 (HSP70)

Answer 23

Chaperone protein that protects unfolded proteins.

Question 24

Mann 6-P signal

Answer 24

To the lysosome (SP).

Question 25

Trp-rich domain signal

Answer 25

To a secretory vesicle (SP).

Question 26

N terminal hydrophobic a-helix signal

Answer 26

To the mitochondria (CP).

Question 27

K and R rich signal

Answer 27

To the nucleus (CP).

Question 28

SKL signal

Answer 28

To peroxisome (CP).

Question 29

KDEL signal

Answer 29

To the ER lumen (SP).

Question 30

N terminal nonpolar region signal

Answer 30

To the cell membrane - not for vesicular secretion (SP).

Question 31

Two properties of secretory pathway proteins

Answer 31

1. 1-2 basic AAs near the N terminus (R, K).

2. Extremely hydrophobic sequence (10-15 residues) on the C terminus side of the basic AAs.

Question 32

Signal recognition particle (SRP)

Answer 32

Binds ot the ER-targeted signal and the ribosome during translation. SRP wraps around ribosome-mRNA-peptide complex halting translation temporarily (SP).

Question 33

I-cell disease

Answer 33

Lysosomal storage disease. Mann 6-P tagging is defective. Failure to thrive, developmental delays, abnormal skeleton, hepatomegaly, recurrent RTIs, death by 7.

Question 34

Protein folding

Answer 34

Small proteins can fold spontaneously, but large proteins require chaperones.

Question 35

Chaperonins

Answer 35

Catalyze folding of proteins using ATP.

Question 36

Proteolytic cleavage

Answer 36

Converts inactive forms of enzymes to active forms (e.g. proisulin to insulin).

Question 37

Covalent modifications (4)

Answer 37

1. Glycosylation
2. Phosphorylation
3. Disulfide bond formation
4. Acetylation

Question 38

Glycosylation

Answer 38

Extracellular proteins are glycosylated (glycoproteins) via O-linked or N-linked.

Question 39

O-linked glycolsylation

Answer 39

Formed with OH- groups of S, T.

Question 40

N-linked glycosylation

Answer 40

Formed with N always.

Question 41

Phosphorylation

Answer 41

Turns on/off proteins. Phosphatases/kinases add or subtract phosphates.

Question 42

Disulfide bonds

Answer 42

Can be inter or intra molecular to stabilize proteins via a thiol groups of 2 Cys. Occurs in ER lumen and is facilitated by protein disulfide isomerases.

Question 43

Acetylation

Answer 43

Typically occurs on K residues and use ACoA as acetyl group donor. Histones commonly acetylated and catalyzed by HAT or HDAC enzymes. Histone acetylation patterns are inheritable.

Question 44

Post-translational modifications of collagen

Answer 44

Done via lysyl and prolyl hydroxylases, which requires Vit C. Defects result in skin, bone, joint disorders.

Question 45

Alzheimer’s Disease

Answer 45

APP breaks down to form amyloid beta peptide (AB). Misfolding of AB forms plaques in brain (extracellular) and hyperphosphorylation of Tau (intracellular).

Question 46

Parkinson’s Disease

Answer 46

Aggregation of a-synclein (AS) protein deposit as Lewy bodies in dopaminergic neurons of the substantia negra, causing death of these neurons.

Question 47

Huntington’s Disease

Answer 47

Mutation of Huntington gene causing expansion of CAG triplets repeats which results in polyglutamine repeat. Selective death of cells in basal ganglia causes the symptoms.

Question 48

Creutzfeldt-Jakob Disease

Answer 48

Caused by misfolding of prions. It is transmissable, as it converts normally folded proteins to misfolded ones. Belongs to TSEs (transmissable spongiform ecephalopathies).