Protein Ligand Binding Flashcards
(79 cards)
1
Q
What 4 characteristics affect protein structure?
A
- size, shape, charge + polarity
2
Q
What is a stable interaction
A
- prosthetic group permanently associated with a protein + is required for protein function
3
Q
What is transient interaction?
A
Ligand that is bound reversible to a protein
4
Q
What does induced fit mean?
A
- structural adaptation between protein, ligand or both
- conformational change can make the binding site more complementary
5
Q
What is the purpose of the catalytic/active site?
A
- binds substrate + facilitates chemical transformation
- enzyme changes bound substrate
6
Q
What are the 4 key concepts of protein-ligand interactions?
A
- reversible binding
- specificity
- conformational change
- regulation
7
Q
Why is regulation required for non-enzymatic proteins?
A
- controls binding protein affinity
- modulate protein-mediated transport
8
Q
What are 2 characteristics of oxygen?
A
- poorly soluble in aqueous solutions
- inefficient diffusion through tissues
9
Q
What is the importance of oxygen binding to transition metals?
A
Gives oxygen binding proteins SPECIFICITY
10
Q
What does free iron promote the formation of? What decreases this process?
A
- ROS
- decreased when iron added to heme
11
Q
True or False: Fe2+ (ferrous) binds oxygen reversibly.
A
True
12
Q
Why does Fe3+ not bind oxygen?
A
2 oxygen can irreversibly oxidize Fe2+ into Fe3+
13
Q
What substances bind to the final 2 coordination bonds in heme?
A
oxygen + histidine
14
Q
Describe characteristics of myoglobin.
A
- monomer
- binds + stores oxygen in muscle
15
Q
Describe characteristics of hemoglobin.
A
- tetramer –> 2 alpha + 2 beta globins
- oxygen transporter in blood
16
Q
Describe characteristics of leghemoglobin.
A
- sequesters oxygen
- protects oxygen-sensitive enzymes in nitrogen-fixing bacteria
17
Q
What is the polarity surrounding the heme-binding pocket?
A
heme surrounded by non-polar residues except for two histidine (polar)
18
Q
What is the proximal histidine? What does it represent? Where is it located?
A
F8 –> 8th AA on alpha helix F
located at 5th coordination bond
19
Q
What is the distal histidine? Where is it located? What is it’s function?
A
E7 - 7th AA on alpha-helix E
- close but not directly bonded to heme
- gives additional specificity to oxygen-binding proteins
20
Q
Where does oxygen bind on heme?
A
6th coordination bond
21
Q
True or False: Proximal + distal histidine residues are different in myoglobin, alpha-hemoglobin + beta-hemoglobin.
A
False: Proximal + distal histidine residues are maintained (same) in myoglobin, alpha-hemoglobin + beta-hemoglobin.
22
Q
Binding of oxygen to myoglobin depends on what?
A
- structure around ligand-binding site
- flexibility of protein
23
Q
What is breathing?
A
small molecular motions of AA side chains on nanosecond time scale
24
Q
What causes strong absorption of visible light in heme?
A
conjugated double bonds
25
What colour is oxy-heme?
Red --> reflects red + absorbs blue light
26
What colour is deoxy-heme?
Blue --> reflects blue + absorbs red
27
What does a high Kd value indicate?
weak interactions between ligand + protein
28
What does a low Kd value indicate?
strong interaction between ligand + protein
29
What happens if the concentration of ligand-bound protein increases [PL]?
Ligand concentration [L] and free protein concentration [P] decrease
30
What does P50 refer to when talking about oxygen binding proteins?
partial pressure of oxygen at which half the ligand-binding sites are occupied
31
Why does CO bind to free heme (not bound to globin) instead of oxygen?
has a higher affinity than oxygen, therefore oxygen cannot bind
32
What is the difference between oxygen binding to heme + CO binding to heme?
[- oxygen binds at an angle
- CO binds heme upright
33
Does Mb have a high or low affinity for oxygen? Does pO2 affect Mb?
- high affinity for oxygen
- not affected by pO2 as Mb becomes fully saturated past a certain point
34
Why does hemoglobin bind oxygen for transport?
- high affinity for oxygen in lungs
- low affinity for oxygen allowing release to tissues
35
Why does Mb not transport oxygen?
Mb becomes completely saturated + has a strong affinity for oxygen
Mb would not release oxygen
Mb used to store oxygen
36
What are the strongest interfaces in hemoglobin?
alpha 1 - beta 1 + alpha 2 - beta 2
37
What kind of interactions hold together hemoglobin subunits? List based on weakest to strongest interaction.
Hydrophobic Interaction
H-bonds
Ion Pairs
38
What is the purpose of ion pairs (salt bridges)?
stabilize the conformation of hemoglobin
39
What subunits form salt bridges in hemoglobin?
Beta-1 subunit His HC3 forms salt bridges with Beta-1 Asp FG1 + Alpha-2 Lys C5
40
Why is His HC3 able to bind to Lys C5?
His HC3 has (-) COO at C-terminal allowing it to bond to (+) Lys C5
41
What is the partial oxygen level in lungs?
13 kPa
42
What is the partial oxygen level in peripheral tissues?
4 kPa
43
How does hemoglobin release oxygen into tissues?
undergoes conformation change from high affinity state (R state) into low affinity state (T state)
44
What is allosteric protein? Does it cause an increase or decrease in binding affinity?
Cooperative binding --> binding of a ligand to one site affects the binding proteins of another site
can cause either increase or decrease in binding affinity
45
Explain hemoglobin allostery.
- oxygen binds to one subunit of Hb + alters affinity for oxygen in adjacent subunits
- induces conformational change pushing adjacent subunits into R state
46
What is a homotropic modulator?
ligand + modulator = identical
47
What is a heterotropic modulator?
ligand + modulator = different
48
Is oxygen a homotropic or heterotropic modulator?
Activating homotropic modulator (increases affinity)
49
Describe the change in Hb shape when oxygen binds.
- T state to R state
- repositioning of Fe2+ in heme reduces bend in porphyrin ring
- His F8 pulls back causing shift in helix F position
- ion pair formed by His HC3 broken
50
What happens to the ion pairs in deoxy-hemoglobin? What state is it in?
- ion pair between Beta-1 His HC3 + Beta 1 Asp FG1 + alpha Lys C5
- T state
51
What happens to the ion pairs in oxy-hemoglobin? What state is it in?
- R state
- His HC3 is not participating in salt bridges (still some salt bridges present)
52
What causes the T state to R state transition?
oxygen binds to Hb
53
True or False: Hemoglobin releases about 37% of its total capacity of oxygen into tissues.
True
54
What does a sigmoidal curve represent?
Cooperative binding
55
What is the pO2 in lungs? Does Hb bind or release oxygen?
13 kPa + binds oxygen (R state)
56
What is pO2 in tissues? Does Hb bind or release oxygen?
4 kPa + releases oxygen (T state)
57
What does nH < 1 represent in Hill's equation?
negative cooperativity (ligand binding on one site decreases affinity at other sites)
58
What does nH = 1 represent in Hill's equation?
no cooperativity (ligand binding on one site has no affect at other sites)
59
What does nH > 1 represent in Hill's equation?
positive cooperativity (ligand binding on one site increases affinity at other sites)
60
How many conformations can exist in the concerted model? Are subunits dependent or independent? Explain.
- 2 conformations (subunits can exist in more than 1 conformation)
- subunits are dependent (change conformation simultaneously)
61
Are subunits dependent or independent in the sequential model?
- Independent (subunits don't have to all change conformation at same time)
- conformation change in one subunit promotes conformational change in adjacent subunit
62
What 2 things are required for cooperative binding?
- more than 1 ligand-binding site
- interaction between ligand-binding sites
63
Explain anemia vs. CO poisoning. Which is worse?
- CO binds to Hb and changes the affinity for oxygen
- Oxygen doesn't get released into tissues
- Anemic individuals bind less oxygen than normal Hb individuals
- release only about 15% of oxygen into tissues
- CO is worse as less oxygen gets released into tissues (approx. 5%)
64
What is the relationship between H+ and CO2 in terms of oxygen bind to Hb? Explain.
- negative heterotropic allostery
- H+ and CO2 decreases affinity of O2 in Hb
65
Describe the situation in peripheral tissues in terms of pH and affinity for oxygen in Hb? What state is Hb in?
- pH is low (high H+) and CO2 is high (metabolism)
- Hb binds H+ and CO2
- oxygen affinity decreases
- T state
66
Describe the situation in the capillary of the lungs in terms of pH and affinity for oxygen in Hb? What state is Hb in?
- Hb releases CO2 + H+
- CO2 low + pH rises (low H+) --> excreted by exhalation
- oxygen affinity increases
- R state
67
What does H+ bind in Hb? What does this formation do?
- His HC3
- favours formation of ion pair (His HC3 + Asp FG1)
- stabilizes T state (more salt bridges)
- pH drops (T state) = decrease in oxygen affinity
68
Explain the reaction between hemoglobin and CO2.
-CO2 reacts with amino group + forms carbaminohemoglobin
- carbamate group participates in salt bridges + stabilizes T state
- produces H+ (pH decreases) = Bohr effect
69
What is the function of BPG? Where does it bind?
- forms salt bridges in beta subunits
- stabilizes T state + decreases oxygen affinity in Hb
- binds to central cavity in heme
70
What kind of modulator is BPG?
- heterotropic allosteric modulator
71
What state does BPG bind in?
- T state (deoxy Hb)
72
Explain the binding purpose of BPG? What would happen if there was no BPG?
- binding of oxygen to Hb pushes Hb into R state (BPG released)
- BPG in low oxygen environments helps push Hb to release O2 (lowers Hb affinity for O2)
- No BPG = Hb would barely release any oxygen into tissues (oxygen would not bind)
73
Explain the BPG role in oxygen transfer between maternal and fetal blood.
-HbFetus has low affinity for BPG (high affinity for oxygen - wants to bind oxygen)
- Kd for BPG is lower in HbAdult than fetus (low affinity for oxygen - wants to release oxygen)
- Kd for oxygen is lower in HbFetus than HbAdult (stronger interaction between Hb + oxygen in fetus)
- Fetus takes oxygen from maternal Hb in tissues (4 kPa)
74
True or False: Binding of BPG affects binding of oxygen as they bind at the same site.
False: Binding of BPG affects binding of oxygen, even though they bind at DIFFERENT sites.
75
True or False: Hb is capable of perceiving information from its environment.
True
76
What AA gets replaced in sickle-cell anemia? Is this AA hydrophobic or hydrophilic?
- Glutamate A3 on beta-subunit gets changed to a Valine
- Val = hydrophobic
77
What happens to the HbSickle Cell tetramers?
- tetramers have 2 fewer negative AA
- deoxy-Hb (T state) tetramers aggregate (proteins cluster together + form large structures due to abnormal hydrophobic interactions)
78
Explain what can happen to RBC containing HbSickle Cell.
- HbSickle Cell fibers are fragile
- capillaries get clogged, burst open, + lyse
- fewer RBC = less Hb = anemia
79
Explain how HbSickle Cell carriers have immunity to malaria?
- one copy of gene of HbSickle Cell (person has normal RBC morphologically)
- person infected with malaria induces the "sickling" of RBC
- body not used to sickling cell + immune system views sickled cell as "foreign substance"
- infected cell (sickled cell + parasite) is removed from the body
