Lecture 7 Flashcards
What is the Chou-Fasman method?
an empirical technique for the prediction of secondary structures in proteins
Define Tertiary Strucutre
Tertiary Structure: spatial arrangement of AA residues that are far apart in the sequence an to the pattern of disulfide (S-S) bonds
Give an example of a protein with tertiary structure
myoglobin
State the function of Protein disulfide isomerase (PDI)
Protein disulfide isomerase (PDI) rearranges the polypeptide’s non-native S-S bonds into “correct” disulfide bonds
(this assumes that the disulfide bond that is present prior to the action of PDI is an “incorrect” disulfide bond (incorrectly folded))
Protein Folding is a highley ______ process
cooperative
Compare extracellular and intracellular proteins in terms of the amount of S-S bonds they feature
Extracellular proteins often have several S-S bonds
Intracellular proteins usually lack S-S bonds
The protein folding and unfolding process is an “all or none” process. explain why
The molten globule stage that initiates folding is an intermediate stage that is very short
When proteins are denatured, what level of organization is the first to be disassembled?
the tertiary structure
What happens if there is only a partial loss of folding in a protein?
even a partial loss of folding structure will destabilize the remainder of the protein folding structure
What is Quaternary Structure? give an example of a protein with quaternary structure?
Quaternary Structure: spatial arrangement of subunits and the nature of their interactions
The alpha2Beta2 tetramer of human hemoglobin
State the 3 groups of Accessory proteins
PDI (Protein disulfide isomerases)
PPI (peptidyl prolyl cis-trans isomerases)
Molecular Chaperones (variety of molecules that serve to protect peptide structures)
Describe what drives HSP 70 (HSP 40) and it’s function
ATP-driven
reverses misfolds ; newly synthesized proteins ; unfold/refold of trafficked proteins
What is HSP 90 mainly used for?
the protection of signal transduction proteins
What is significant about Nucleoplasmins?
they were the first molecular chaperone to be discovered
Mitochondria contain their own heat shock protein molecules that are distinct from those that function in the cytosol. State these 2 HSP’s
HSP60 and HSP70
State the 3 conditions and the 3 chemicals that are known to denature proteins
Conditions:
Heat
pH (extremes)
Agitation
Chemicals:
Detergents (SDS)
Chaotropic Agents (urea, guanidine hydrochloride)
Organic Solvents (TCA) [trichloroacetic acid]
State the 5 methods of analysis that are used to determine if proteins are denatured or not
- Turbidity
- Circular Dichroism (CD)
- UV Absorption
- Fluorescence
- Biological Activity
What exactly is CD (Circular Dichroism) measuring in order to determine if proteins are denatured or not?
CD measures the differential absorption of right-handed and left-handed circularly polarized light resulting from the molecular asymmetry involving as chromophore group
(it is basically used to study the conformation of proteins in solution)
What are molecular chaperones?
Molecular chaperones: essential proteins that bind to unfolded and partially folded polypeptide chains in order to prevent the improper association of exposed hydrophobic segments
True or False:
Non-native folding, polypeptide aggregation, and precipitation are the only type of protein changes that can occur in the presence of molecular chaperones. explain.
False.
Non-native folding, polypeptide aggregation, and precipitation WILL NOT occur in the presence of molecular chaperones, preventing these is the main function of molecular chaperones
Molecular chaperones allow misfolded proteins to refold into their ____ _____.
Native Conformation
State the 2 major classes of Molecular chaperones and describe their functions
HSP’s: proteins that help repair any denaturation that may occur to a protein that is exposed to heat
Chaperonins: a protein folding container)
Describe the 2 main HSP’s (describe them)
HSP70 family: coordinates cellular function by directing substrates for unfolding, disaggregation, refolding or degradation
HSP90 family: Integrates signalling functions, acting at a stage of folding of substrates
State the 2 main Chaperonins
HSP60 proteins (GroEL)
HSP10 proteins (GroES) (Co-chaperone)
Describe the structure of a chaperonin and outline the process by which it folds proteins
Chaperonins are a hollow cylinder with a cap on the top
(looks like a washing machine)
The protein enters, the cap closes and the cylindrical body changes shape to create a hydrophilic environment for the protein inside
The protein then folds, then the cap comes off, and a folded protein exits
A number of pathological diseases are associated with protein aggregation. This is why ____ _____ is so tightly regulated
Protein turnover
state the 2 main reasons why a cell would need to remove a protein
The protein is too old
The protein is damaged
Name the main player in the protein homeostasis system that is known for degrading ubiquitinated proteins
proteasomes (a large protease complex)
Describe the structure of the 26S proteasome
a 20S catalytic core with 2 19S regulatory units on the top and bottom of it
Describe the functions of the 19S regulatory unit of the 26S proteasome (3 of them)
- recognizes ubiquitinated proteins
- isopeptidase cleaves off the ubiquitin
- directs the protein into the catalytics core (20s)
Describe the 20S regulatory unit of the 26S proteasome
It is a catalytic core composed of 28 subunits that form a “sealed barrel”
(access is controlled by 19S regulatory subunits on either side of it)
Describe the process by which proteasomes and other proteases create free AA’s from ubiquitinated proteins.
Ubiquitinated proteins are processed into peptide fragments
peptide fragments are then further digested into free AA’s that can now be used for other biosynthetic reactions (recycling)
