Lecture 10 Flashcards
What are some of the features of the serine protease catalytic mechanism that contributes to lowering of the activation nrg. Contributes to inc in rate of rxn?
Enzyme catalysis (Serine Protease Mechanism)
- Binding site (binds substrate in specific way. Inc rate by increase local effective concentration). Maybe 2 substrates have to come together and keeping them there long enough for them to react. Having them in local effective conc can help guide fxn forward. Important for specificity. Enzymes don’t only inc catalytic rate but eliminate a lot of the side products u can get. Specificity for substrate. What kind of features of the substrate are recognized? Electrical stat/ shape (steric)/ stereochemistry/ polar
- Orbital orientation (getting reactants to be in right juxtaposition). The 1st stage of the rxn in the orientation of the nucleophile the 2nd stage is orientation of second nucleophile (the water). (Ser/H2O). Certain angle, origination, and distance that is important for the nucleophilic attack. This is called the Burgi angle (angle of attack on carbonyl of nucleophile).
- Shifting of pKa values. (e.g. weird enzyme not as histidine as general base
- Electrostatic stabilization (TS stabilization)
- Covalent catalysis (not all enzymes have this but directly brings substrate to different pathway)
What does the hill coefficient (n) describe?
Cooperativity; it describes the fraction of the macromolecule saturated by ligand as a fxn of ligand conc
What does
n = 1
n > 1
n < 1
mean?
- n=1 (no cooperativity) completely independent binding, regardless of how many additional ligands are already bound
- n>1 (+ cooperativity) Once one LIGAND molecule is BOUND to the PROTEIN, its AFFINITY for other ligand molecules INCREASES.
- n<1 ( - cooperativity) Once one ligand molecule is bound to the LIGAND, its affinity for other ligand molecules DECREASES.
How is myoglobin structurally related and functionally related? How do they differ in terms of cooperativity
Myoglobin - monomer, no cooperativity
Hemoglobin - oligomer, cooperativity
Functionally related cause bind oxygen
What is the eqn for the reversible binding of O2 to myoglobin?
Mb(O2) —->
What is Y for Mb?
Fractional saturation of Mb w/O2
Eqn for Y for Mb?
Y = [MbO2] / [MbO2] + [Mb]
Draw Y vs. pO2 plot and hill plot
Slide 4
What is the eqn for the reversible binding of O2 to the 4 sites of Hb?
Hb(O2)4 —->
What is Y?
Fractional saturation of Hb w/O2
What is eqn for Y for Hb?
Y = [Hb(O2)4] / [Hb(O2)4 + [Hb]
Draw Y plot for Hb and Hill plot
Slide 13
Organic part of Heme?
____________
What type of iron?
____________
What type of cofactor?
_____________
Geometry of coordination? \_\_\_\_\_\_\_\_\_\_ \_\_\_\_\_\_\_\_\_\_ \_\_\_\_\_\_\_\_\_\_ \_\_\_\_\_\_\_\_\_\_
Organic part of Heme?
1. Porphyrin ring
What type of iron? Fe II (Fe2+) (Ferrous)
What type of cofactor?
Prosthetic group
Geometry of coordination?
- Octahedral
- Tetragonal bipyramidal
- 5 N ligands
- 1 O ligand
Slide 7
N/A
Compare Mb vs. Hb in terms of oxygen saturation
Slide 8
Myoglobin
- Fxn?
- Found where?
- How does O2 get in and out
- Fxns to store O2
- Found in muscle cells
- Gives meat its red colour, when exposed to O2
- deoxymyoglobin is PURPLE - O2 gets in and out due to breathing (dynamics) of the protein
What is the most abundant protein in mammals?
Hb
Where is Hb located?
Erythrocyte cytosol
Describe the 3-D structure of Hb
- 2 α/β protomers
- A dimer of heterodimers
- 4 heme, 4 O2 binding sites
- Globin fold (globular) – all α protein fold
Describe structure of deoxyHb
- Water binds to Fe
2. Iron atom lays BELOW the PLANE of the HEME.
How does the structure of Hb change when going from deoxyHb to oxyHb
In deoxyHb, water binds to Fe. The Fe atom lays below the plane of the heme.
In oxyHb, the binding of O2 pulls Fe into plane of the heme. This pulls on the PROXIMAL His, which pulls out of helix. This motion is then propagated thru the protein, leading to cooperative O2 binding.
The cooperativity is achieved through domain rotation ~ ____ degree in _____ rotation
The cooperativity is achieved through domain rotation ~ 15 degree in protamer rotation
What are the 4 other molecules that can bind to Hb besides O2?
- nitric oxide (NO): cardiovascular signalling molecule.
- carbon monoxide (CO): poison. Forms a stable complex hard to remove
- carbon dioxide (CO2): product of respiration
- 2,3-bisphosphoglycerate (2,3-BPG): binds to deoxygenated Hb promotes release of O2, tense state
Slide 19
N/A
What is the molecule called when CO2 binds to Hb
Carbaminohemoglobin
How many molecules can Hb bind to of Co2?
- Hemoglobin can bind to 4 molecules of CO2
- The CO2 molecules form a carbamate with the 4 terminal-amine groups of the 4 protein chains in the deoxy form of the molecule.
Draw eqn for carbamate molecule
Protein-NH2 + CO2
—->
- Where does 2,3-BPG bind to Hb?
- How does this effect the affinity for O2?
- How many molecule(s) of 2,3-BPG bind per Hb?
- Binds to beta-subunits
Near center - Stabilizes T state
reduces oxygen affinity - 1 molecule binds per Hb
2,3-BPG causes a _____ shift in the Hg-oxygen saturation curve
RIGHT
