Lecture 3 Flashcards
Types of post translational modifications?
- Cleavage by peptidases
- Covalent modification of N-terminus
- Covalent modification of side chains
Types of side chain modifcations?
-Phosphorylation
-Methylation
-Acetylation
-Glycosylation
-Ubiquitination
Result of side chain modifications?
-Change the conformation/surface of a protein and affect the proteins it can bind/block
Are side chain modifications reversib le?
Yes
Side chain modification are mediated by…?
Enzymes
Major side chain modification?
Phosphorylation
Phosphorylation + amino acids?
-Changes the charge + size of a.a
-Hydroxyl groups are phosphorylated
-Amino acids (Tyrosine, Threonine, Serine)
Kinases?
Transfer phosphates from ATP
-Specific(only specific kinases work for specific side chains and peptide sequences
Phosphatase?
Remove phosphate
Two types of kinases?
-Ser/Thr kinases
-Tyr kinases
Why are serine and threonine able to be phosphorylated by the same kinases?
Have similar structures
Phosphatases?
Two types
- Ser/Thr phosphatases
-Tyr phsophatases
What is a phosphomimic?
Researchers have developped a way to recreate the phosphorylation of Thr, Ser and Thr
How do phosphomimics work?
Substitute serine with aspartic acid , aspartic acid has a similar strucutre to phosphorylated serine
Phosphomimic if we don’t want serine to be phosphorylated?
Substitue serine for alanine which does not contain a hydroxyl and cannot be phosphorylated
Half-life and phosphorylation?
Phosphorylation is often a signal prior to degradation of a protein and therefore can be an indicator of a proteins half life. Phosphorylation can target proteins for ubiquitination
What side chains are acetylated?
Lysines
Lysine acetyltransferases (KATs)?
Add the acetly group to a lysine side chan
Lysine deacetylases(KDACs) ?
Deacetylate the lysines
HATs(histone acetyl transferases)?
Proteins in DNA that are acetylated
How does the acetyl group affect the lysine?
-no longer has a positive charge(no more ionic interactions)
-Changes the polarity
Histone acetylation?
Prior to acetylation lysines on histones are positive and able to interact with the negatively charged DNA which will pack the DNA preventing transcription
Post acetylation histones can no longer interact and compact DNA which results in active transcription
Methylation
-Lysine + Arginine
-Increases the size of the side chains
Methylarginines?
Add methyl groups to arginines
lysine methyltransferases(KMTs) vs Lysine demethylases(KDMs)
KMTs: Add methyl group to lysine
KDMs: remove methyl group from lysine
T/F: Acetylation, phosphorylation and methylation provide new binding sites for proteins?
True
Native state?
Completely folded conformation of a protein in the active protein conformation
Native state depends on?
Native structure is determined by the primary sequence
The hydrophobic interactions in the interior of the protein, the polar side chains are in the exterior interacting with the aqueous environment
When do post translation modifications occur?
After the protein has been folded to the native state
T/F: Proteins are soluble?
True, when proteins are properly folded. Since the polar side chains at the exterior of the protein are able to interact with the environment
Delta G of the native structure?
Folding of proteins is spontaneous but in reality is assisted by different biological mechanisms
Folding process?
Folding proceeds through intermediates that have increasing structure to the native state which is the least flexible and most stable structure
Cofactors and native state?
Some proteins require a cofactor to stabalize the native state
Folding Intermediates?
-Have some secondary structures
-Some hydrophobic side chains exposed
-More flexible and disordered
Aggregation ?
Hydrophobic regions prefer to interact with other hydrophobic regions. This means unfolded proteins aggregate leading to insolubility.
Why are aggregates so bad?
They have extremely low gibbs free energy which means they are not favourably undone.
When do we find Protein misfolding?
- Immediately after protein synthesis (protein has not been fully folded yet and hydrophobic residues are exposed)
-Ligand not available (no co factor)
-Genetic mutation
-Harmful environmental conditions(ex. heat)
-Aging (decrease in the efficiency of the protein quality control mechanism)
Neurodegeneration?
Proteins tend to aggregate due to exposed hydrophobic regions which leads to diseases such as Alzheimers, Parkinsons, ALS and dementia
Mutations?
Changes in the polypeptide sequence either insertion, deletion, substitution or premature stop codon
Mutation substitution to similar amino acid vs very different?
Similar: could have little to no effect on the protein
Different: Could disrupt folding/function of the protein
Protein Homeostasis?
Network of components that act to maintain proteins in the correct concentrations, conformation, and subcellular location
Chaperones?
Bind to hydrophobic regions of proteins and assist in folding and prevent protein aggregation
When are chaperones needed?
Chaperones are constantly expressed and essential even under non-stress conditions
How do cells respond to stress that causes protein misfolding?
By increasing the expression of chaperones
Heat Shock Response (HSR)
-Refers to misfolded proteins in the cytosol/nucleus
-Protects against cell death
Unfolded Protein Response(UPR)?
-Refers to misfolded proteins in the ER
-Can promote cell death if stress if too severe
How were Heat Shock Proteins discovered?
When drosophilas were at extreme temperatures there were specific regions of the chromosome that were always transcriptionally active these were the genes of the heat shock proteins. Under normal conditions, the chromosome expresses completely different proteins
When HSP expressed?
-Marathon running, alcohol, hypoxia
Constitutive vs inducible chaperones?
Constitutive: Chaperones always expressed even under normal condition
Inducible : Induced due to stress conditions
When are unfolded cytosolic proteins present?
-heat stress
-oxidative damage
-proteasome inhibition
HSF1?
Heat shcok factor one mediates the heat shcok factor response and activates the transcription of Heat shock chaperones
Inactive HSF1 vs Active?
Inactive: Monomer(has all the domains but has to form the quarternary structure to be active)
Active: Trimer(quarternary structure)
Heat Shock Element?
sequence in the promoter where the HSF1 binds when it is active
Domains of HSF1?
- DNA binding domain
- Trimerization domain
- Transcription activation domain
