Lecture 8: Protein Structure Flashcards
A molecule that binds to a protein is called?
A ligand
A region in the protein where the ligand binds is called
The binding site
What type of bonding interaction do ligands bind?
Noncovalent interactions
What is Ka and Kd. Draw their equations based on a Protein (P) and Ligand (L)
ka [P][L] = kd [PL]
ka = [PL] / [P][L] = 1/kd
kd = [P][L] / [PL]
How can we directly measure [PL]
Ptot = [P] + [PL]
[PL] = Ptot [L] / Kd + [L]
What it the equation for fraction of occupied binding sites?
Or x = [PL] / [PL] + [P]
What is the gibbs free equatino with the dissociation constant
ΔG° = RT ln(kd)
What does a strong binding or weak binding look like with kd
Strong binding: kd < 10 nm
Weak binding: kd > 10 um
The smaller the kd the strong the bond
What is the lock and key model?
Assumes that complementary surfaces of enzymes are preformed
Complementary in
- Size
- Shape
- Charge
- Hydrophobic/philic character
What is induced fit?
Conformational changes may occur upon ligand binding to enzyme and ligand
- allows for tighter binding
- allows for high affinity for different ligands
What is cooperativity?
As the change in properties of binding at one site as a function of whether or not another site is occupied
- Often caused by structural change from binding ligand on one site
- can make other sites stronger or weaker
What is the Hill equation
n = Hill coefficient (the degree of cooperativity)
What biological problems do we have with O2
- Proteins lack affinity for it
- Binding to some transitional metals would create free radical
- Fe3+ is very reactive
What is the biological solution to O2
Capture oxygen molecule with heme that is protein bound
Why is CO better at binding heme than O2
because the carbon in CO has a filled lone electron pair that can be donated to the vacant d-orbitals on the Fe2+
How does O2 binding affect the color or Fe
- Fe2+ without O2 has intense soret band at 429 nm
- With O2 shifts to 414 nm
- Via UV-Vis
- Deoxyhemoglobin appears purple
- oxyhemoglobin appears red
For effective transport, affinity must vary with pO2
What is Tense state and Relaxed state
Tense state
- More interactions, more stable
- lower affinity for O2
Relaxed State
- Fewer interactions, more flexible
- higher affinity for O2
Does O2 cause a T → R change or R → T change?
O2 binding trigger a T → R conformational change
Involves breaking ion pairs between the a1-b2 interface
What is the a1-b2 interface?
What triggers the conformational change in hemoglobin?
By oxygen binding
What effect does pH binding have on hemoglobin
- H+ binds to Hb and stabilizes the T state this protonates His146 which then forms a salt bridge with Asp94
- Leads to the release of O2
- Increases Kd
What is the Bohn effect
The effect pH has on the binding of O2 to hemoglobin. This increases the efficiency of the O2 transport.
What is carbamate?
15-20% of CO2 is exported in the form of a carbamate on the amino terminal residue of each of the polypeptide subunits
It produces an H contributing to Bohr effect
Stabilizes T state
How does 2,3 Bisphosphoglycerate regulate O2? (know structure)
- Produced as intermediate of glycolysis
- Small negatively charged molecule, bind to the positively charged central cavity of Hb
- Stabilizes the T state and allows for better release in Tissue and adaptation to changes in altitude (higher Kd)
What mutation on hemoglobin occurs to form sickle cell
Glutamate-6 to valine
Who are the key players for cellular immunity?
Targets own cells that have been infected
macrophages, killer T cells, and inflammatory T cells
What is humoral fluid immune system
Targets extracellular pathogens
makes soluble antibodies
Keyplayers: B-lymphocytes and helper T cells
How does an antibody look?
Two heavy chains and two light chains
Light: one constant and one variable domain
Heavy: Three constant and one variable domain
Variable Domain: make up the antigen-binding site (two per antibody) and are hypervariable.
How do antigen bind?
Via induced fit
How does ELISA work?
