Post Translational Modifications / Ubiquitination Flashcards
1
Q
What are post translational modifications and examples
A
- Stability, function and activity to ensure gene complexity, changes occur to polypeptide chain before it becomes an active protein Examples - Protein folding - Proteolytic cleavage of proteins - Acetylation - Glycosylation - Methylation - Phosphorylation - Ubiquitin - Targeted protein degradation
2
Q
How is gene expression regulated
A
- Folding of polypeptide into correct 3D structure
- Transport to the correct location
- Post translational processing (phosphorylation and glycosylation)
- Association with other protein co-factors
3
Q
What is the purpose of protein folding
A
- Protein structure assumes its functional shape or conformation
- AA interact with each other to produce a well-defined 3D structure, folded protein, known as the native state
- Polypeptide chains spontaneously fold so that hydrophobic AA are on the inside of the structure
- Spontaneous folding leads to formation of many different structures (most inactive)
- At high concentrations unfolded proteins form insoluble aggregates through interaction of side chains
4
Q
What are chaperones
A
- Help other proteins fold appropriately
- Prevent protein-protein interactions during folding process
- Prevents formation of insoluble aggregates
- HSP: Aid protein folding and reducing formation of insoluble protein aggregates in the cell, important as messengers
5
Q
What is aberrant protein folding
A
- Mis-folding
- Numerous diseases result from incorrectly folded proteins
- Diabetes, obesity, cystic fibrosis, haemophilia A and B, blood coagulation disease, goitre, gaucher’s disease
6
Q
What is the unfolded protein response
A
- Under normal conditions BIP binds to IRE1, ATF6 and PERK
- When unfolded proteins accumulate in ER, BIP is released
- Initiates signal transduction pathways leading to induction of gene expression
- Synthesis of chaperones in increased to decrease conc of unfolded proteins
- If unfolded proteins continue to accumulate apoptosis is induced (CHOP, caspases)
- Can cause translation attenuation which halts progression of protein synthesis in attempt to resolve production of insoluble proteins
7
Q
What occurs after the unfolded protein response
A
- Activated IRE1 recruits TRAF2 to elicit JNK-P phosphorylation, activating caspases
- Activated PERK and ATF6 increase expression of nuclear genes for apoptotic effector CHOP
- Inhibits expression of Bcl2 and induces expression of genes for apoptotic promoting factors Gadd34, Trb3 and Dr5
- Release of Ca from ER leads to mitochondrial generation of ROS, contributes to cell death
8
Q
What is proteolytic cleavage of proteins
A
- Breaking peptide bonds between AA in proteins
- Creation of a stable form
- Some proteins are synthesised as inactive precursors that are activated under proper physiological conditions by proteolysis
- Inactive precursor proteins that are activated by removal of polypeptides are termed pro-proteins
9
Q
What are examples of proteolytic cleavage
A
- Pro-Caspase 3: Will cause apoptosis if it circulates the body in activate form
- Insulin: Preproinsulin (110AA) to proinsulin to insulin (51AA)
- POMC: Undergoes glycosylation, acetylation and proteolytic cleavage to produce ACTH and MSH
- Addisons Disease: Negative effect of POMC, insufficient hormones produced,
10
Q
What is glycosylation
A
- The addition of polysaccharide side chains (sugar) to proteins (glycoprotein)
- For a given protein the pattern of glycosylation differs for different species/groups, species specific
- Determines protein structure, function, stability
- Types: N and O linked glycans
- Variations in glycosylation can lead to disease
11
Q
What is the biological significance of glycosylation
A
- Oligosaccharides may be a tissue-specific marker
- Carbohydrates may alter the polarity and solubility
- Bulkiness and -ve charge of oligosaccharide chain may protect protein from attack by proteolytic enzymes
- Markers and therapy for cancerand autoimmune disease
- More efficient production of pharmaceuticals
12
Q
What are N linked glycans
A
- Covalently attached to Asn residues within a consensus sequence (Asn-Xaa-Ser/Thr)
- Enable prediction of modification sites by protein sequence analysis
- Share common penta-saccharide core recognised by lectins and N-glycanase enzymes (PNGase F)
- Aid visualisation of proteins and enrich mass for spectrometry analysis
13
Q
What are O linked glycans
A
- Comparable tools are lacking for study of proteins bearing O-linked glycans
- Mucin-type
- N-acetylgalactosamine (GalNAc) residue linked to hydroxyl group of Ser or Thr
- Complex biosynthetic origin
- Not installed at a defined consensus motif and presence cannot be accurately predicted based on protein’s primary sequence
14
Q
What is notch1 and how is it affected by glycosylation
A
- Notch 1: Consists of a number of EGF repeats, different types of glycan side chains are found attached to these EGF repeats
- Glycosylation: Receptors lacking O or N glycans are functional, lack of O fucose glycan destroys activity
- Notch Defects: Result in perinatal lethality, defects in somitogenesis, T-cell leukaemia, cancers, and developmental defects
15
Q
What is phosphorylation
A
- Most common protein modification that occurs
- Regulates biological activity of a protein
- Numerous growth factors affected by phosphorylation
- Kinases: Phosphorylate proteins
- Phosphatase’s: Remove phosphates
- Protein Kinases: Catalyse reactions of the following type, ATP + protein to phosphoprotein + ADP
