Post-translational Modifications of Proteins

Question 1

Proteolytic processing

Answer 1

Major form of post translational modification which occurs when a protease cleaves one or more bonds in a target protein to modify its activity

Question 2

Covalent amino acid modifications

Answer 2

Collagen (proline, lysine, vitamin C, cysteine), glycosylation (N- and O-linked), phosphorylation (kinase/phosphatase), acetylation and methylation (histones), carboxylation (glutamate, vitamin K)

Question 3

Cleavage of signal peptide steps

Answer 3

1. Membrane bound proteins or secretory have signal peptide (hydrophobic core at N-terminus) synthesized by ER–protein binds to multi-protein complex called signal recognition particle (SRP) 2. Complex binds to SRP receptor on ER membrane 3. Ribosome transferred to the translocon (complex that helps proteins through membrane) 4. GTP hydrolyzed and signal peptide removed by aspartyl signal peptidase (SPP) once in ER lumen

Question 4

Insulin proteolytic processing steps

Answer 4

Occurs in the cell. 1. Proinsulin synthesized in ER with disulfide bond formation 2. Transported to golgi apparatus 3. packaged into secretory vesicles 4. processed by proteases to form mature insulin (_ and _ chain stay connected by disulfide bonds)

Question 5

Protein glycosylation

Answer 5

Non-template. Requires activated sugar (UDP-glucose), a glycosyl transferase, and an acceptor protein. Sugars covalently linked through O-linkages to serine or threonine or N-linked through asparagine.

Question 6

Steps of protein glycosylation (N-linked)

Answer 6

1. Protein synthesis begins and polypeptide chain extruded into RER 2. Branched ogliosaccharide synthesized on dolichol pyrophosphate (lipid carrier) 3. ogliosaccharide transferred from dolichol to amide N of asparagine residue 4. Trimming of carbohydrate begins as protein moves through RER 5. In golgi apparatus further trimming and addition of monosaccharides

Question 7

Two major classes of N-linked saccharides

Answer 7

1. High mannose oglios-two N-acetylglucosamines with many mannose residues 2. Complex oglios-any number of the other types of saccharides including more than the two N-acetylglucosamines

Question 8

High-mannose vs complex saccharides

Answer 8

Whether high-mannose or complex depends on accessibility to saccharide-modifying enzymes in the golgi. If inaccessible saccharide will probably stay in high-mannose form. If accessible then many mannose residues cleaved and will be further modified to make complex saccharide.

Question 9

Mannose 6-P receptors

Answer 9

N-linked glycoproteins can be phosphorylated in golgi at one or more mannose residues. Mannose 6-P receptors bind to phosphorylated glycoprotein for packaging and transporting to lysosomes

Question 10

L-cell disease

Answer 10

Lysosomal storage disease. Cannot phosphorylate mannose residues leading to inability to target hydrolases to lysosomal compartments.

Question 11

Congenital Disorders of Glycosylation (CDG)

Answer 11

Multisystemic disorders with neurological involvement

Question 12

Acetyl-CoA

Answer 12

Acetyl donor for attaching an acetyl group to the N-terminal amino acid

Question 13

Histone acetylation and methylation

Answer 13

Occurs at lysine residues. Acetylation activates transcription and deacetylation silences. Demethylation activates transcription and methylation silences. Regulates gene transcription.

Question 14

Gamma carboxyglutamate (Gla)

Answer 14

Glutamate side chain modified with second carboxyl group to form gamma carboxyglutamic acid. Requires vitamin K. Dicarboxylic acid side chain bonds Ca2+ ions readily.

Question 15

Gamma carboxyglutamic acid

Answer 15

Formed by glutamate side modified with second carboxyl group. Requires vitamin K.

Question 16

Where are gamma carboxyglutamate (Gla) residues found?

Answer 16

Osteoclacin in bone and in blood clotting proteins like prothrombin.

Question 17

What does vitamin K deficiency cause?

Answer 17

Clotting disorders

Question 18

Coumarin (Dicumarol)

Answer 18

Drug that inhibits vitamin K activity

Question 19

Kinases

Answer 19

Enzymes that phosphorylate proteins

Question 20

Phosphatases

Answer 20

Enzymes that dephosphorylate proteins

Question 21

Which AA are subject to phosphorylation?

Answer 21

Serine, theonine, tyrosine

Question 22

Phosphorylation of glycogen synthase and glycogen phosphorylase

Answer 22

In response to epinephrine phosphorylation of synthase inhibits its activity and phosphorylation of phosphorylase increases activity. Together lead to increased hepatic glucose delivery to blood.

Question 23

What amino acid modification controls signal transduction?

Answer 23

Tyrosine phosphorylation

Question 24

What amino acid modificaton controls transcription?

Answer 24

Histone acetylation and methylation

Question 25

Where does signal peptide cleavage occur?

Answer 25

Endoplasmic reticulum

Question 26

Where does glycosylation occur?

Answer 26

Endoplasmic reticulum and golgi apparatus

Question 27

Where does protein folding occur?

Answer 27

Golgi apparatus with the help of chaperones

Question 28

Proteasome structure

Answer 28

Barrel-like proteins found in cytosol and nucleus. Composed of 7 proteins that catalyze 6 major protease activities. Sites located on interior surface of the rings so target protein must enter cell pore before degraded.

Question 29

Proteasome

Answer 29

Degrades unneccessary or damaged proteins by proteolysis (chemical reaction that breaks peptide bonds)

Question 30

Ubiquitin

Answer 30

Small protein that tags other proteins for degredation. Recognized by the proteasome.

Question 31

Ubiquitin-activating enzyme

Answer 31

Binds ubiquitin to the enzyme via ATP

Question 32

Ubiquitin-conjugating enzyme

Answer 32

Transfers uniquitin intermediate to substrate protein. Ubiquitin-protein ligases covalently attach ubiquitin to an internal lysine

Question 33

Ubiquitin-proteasome degredation steps (3)

Answer 33

1. Ubiquitin-activating enzyme binds ubiquitin to the enzyme via ATP 2. Ubiquitin-conjugating enzyme transfers uniquitin intermediate to substrate protein and ubiquitin-protein ligases covalently attach ubiquitin to an internal lysine 3. Tagged proteins degraded by proteasome complex with release of ubiquitin monomers to be re-used

Question 34

What diseases are associated with ubiquitination?

Answer 34

Cancer-defective degredation of growth promoting proteins or key regulators of cell cycle. Neurogenerative diseases like Parkinson’s, Alzheimer’s, ALS where there is a build-up of proteins that should have been degraded and form plaques.

Question 35

SUMO-ylation

Answer 35

Post-translational modification of proteins attaching SUMO (small ubiquitin-like modifier) proteins to target protein. Similar to ubiquitin, directed by enzymatic cascade. Does NOT target proteins for degredation. May act as recognition structure for other proteins to interact with SUMO-ylated protein.