Post-translational Modifications of Proteins Flashcards
Proteolytic processing
Major form of post translational modification which occurs when a protease cleaves one or more bonds in a target protein to modify its activity
Covalent amino acid modifications
Collagen (proline, lysine, vitamin C, cysteine), glycosylation (N- and O-linked), phosphorylation (kinase/phosphatase), acetylation and methylation (histones), carboxylation (glutamate, vitamin K)
Cleavage of signal peptide steps
- 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
Insulin proteolytic processing steps
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)
Protein glycosylation
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.
Steps of protein glycosylation (N-linked)
- 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
Two major classes of N-linked saccharides
- 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
High-mannose vs complex saccharides
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.
Mannose 6-P receptors
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
L-cell disease
Lysosomal storage disease. Cannot phosphorylate mannose residues leading to inability to target hydrolases to lysosomal compartments.
Congenital Disorders of Glycosylation (CDG)
Multisystemic disorders with neurological involvement
Acetyl-CoA
Acetyl donor for attaching an acetyl group to the N-terminal amino acid
Histone acetylation and methylation
Occurs at lysine residues. Acetylation activates transcription and deacetylation silences. Demethylation activates transcription and methylation silences. Regulates gene transcription.
Gamma carboxyglutamate (Gla)
Glutamate side chain modified with second carboxyl group to form gamma carboxyglutamic acid. Requires vitamin K. Dicarboxylic acid side chain bonds Ca2+ ions readily.
Gamma carboxyglutamic acid
Formed by glutamate side modified with second carboxyl group. Requires vitamin K.
Where are gamma carboxyglutamate (Gla) residues found?
Osteoclacin in bone and in blood clotting proteins like prothrombin.
What does vitamin K deficiency cause?
Clotting disorders
Coumarin (Dicumarol)
Drug that inhibits vitamin K activity
Kinases
Enzymes that phosphorylate proteins
Phosphatases
Enzymes that dephosphorylate proteins
Which AA are subject to phosphorylation?
Serine, theonine, tyrosine
Phosphorylation of glycogen synthase and glycogen phosphorylase
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.
What amino acid modification controls signal transduction?
Tyrosine phosphorylation
What amino acid modificaton controls transcription?
Histone acetylation and methylation
Where does signal peptide cleavage occur?
Endoplasmic reticulum
Where does glycosylation occur?
Endoplasmic reticulum and golgi apparatus
Where does protein folding occur?
Golgi apparatus with the help of chaperones
Proteasome structure
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.
Proteasome
Degrades unneccessary or damaged proteins by proteolysis (chemical reaction that breaks peptide bonds)
Ubiquitin
Small protein that tags other proteins for degredation. Recognized by the proteasome.
Ubiquitin-activating enzyme
Binds ubiquitin to the enzyme via ATP
Ubiquitin-conjugating enzyme
Transfers uniquitin intermediate to substrate protein. Ubiquitin-protein ligases covalently attach ubiquitin to an internal lysine
Ubiquitin-proteasome degredation steps (3)
- 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
What diseases are associated with ubiquitination?
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.
SUMO-ylation
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.
