7. Myoglobin and molecular binding Flashcards
T or F: O2 is poorly soluble in aqueous solutions
true
what is the role of myoglobin
functions to both store oxygen and facilitate oxygen diffusion in rapidly contracting muscle tissue
what animal is myoglobin very prevalent in and why
sperm whales, because they need to be under the water for up to 2 hours at a time
how much more myoglobin do sperm whales have than humans
about 30x more
how many polypeptides is myoglobin
1
how many amino acids is myoglobin
153
what is the molecular weight of myoglobin
Mr=16,700
what is the main secondary structure of myoglobin. What percentage is this
78% of residues are in alpha helices
how many helices make up myoglobin
8
what type of amino acids make up the majority of the interior of myoglobin
hydrophobic residues
which hydrophobic residues make up the interior of myoglobin
Valine, Leucine, Isoleucine, Phenylalanine, and methionine
VMILF
what part of myoglobin binds O2
the associated heme group
describe the structure of the heme group of myoglobin
it’s a flat porphyrin ring with a single iron atom in the center. The iron is between 4 nitrogen molcules in the ring, and one atom binds above and one below
why is muscle and blood dark red
heme absorbs light at that wavelength
how are the helices in myoglobin named
A-H
how are the bands connecting the helices named
named after the helices they connect: ie AB, BC, CD, etc
how are residues in the helices named
named for their position in a given helix
ie His F8 is in helix F in the 8th position
what residue do many of the helices end with? how many helices end with this
4 of 8 helices end with proline
why do half of the helices end in proline
they break helices because they don’t fit
in regards to the helices, where is the heme group
it sits in a hydrophobic myoglobin pocket formed by 3 of the 8 helices, shielding it from solvent
what happens to the heme when it’s outside of the pocket
Fe2+ is almost instantly oxidized to Fe3+, which cannot bind O2
which conformation of iron cannot bind O2. When do we see this confirmation
Fe3+ cannot bind O2, and we get this when the heme group is outside of the pocket
what binds to the iron atom in the center of the heme group
N atom (from histidine) binds perpendicular to the plane of the ring, and then O2 binds as well
which histidine does the N come from that binds with the heme group
His F8, aka His residue #93, aka proximal histidine
what happens to O2 when it’s bound to heme
it has a partial negative charge due to its interaction with Fe2+ (this isn’t normal btw)
how does O2 with the negative charge become stabilized
stabilized electrostatically by His64 aka His E7, aka distal his
where does the hydrogen bond form in regards to proximal and distal his and the heme group
hydrogen bond forms between O2 and an N-H on the distal His side chain
what expression is used to represent the reversible binding of a protein to a ligand
P + L ⇌ PL
what is the equilibrium constant for proteins binding to ligands
Ka
what is the expression for Ka (ie how is Ka expressed)
Ka = [PL]/[P][L]
products over reactants
what does Ka represent
the affinity of the ligand to the protein
what does a high Ka represent
high affinity
what are the units of Ka = [PL]/[P][L]
M^-1
what variable represents the rate of the forward reaction
ka
what variable represents the rate of the reverse reaction
kd
how do Ka, ka, and kd relate
Ka = ka/kd
define first order reaction
when there’s one reactant
define second order reaction
when there’s two reactants
what are the units for the rate constant in a first order reaction
s^-1
what are the units for the rate constant in a second order reaction
(M^-1)(s^-1)
T or F: the more ligand we have, the more ligand can bind
true
how do we graphically represent ligand binding
we consider the fractional saturation of the protein with the ligand
= binding sites occupied/total binding sites
what does Y equal
Y=fraction saturation= binding sites occupied/total binding sites = [PL]/([PL]+[P])
what can the Y equation be simplified to
Y = [L]/([L]+1/Ka)
at what value of Y does 1/Ka=[L] on the graph
Y=0.5 corresponds to the [L] that’s equal to 1/Ka
what is Kd
the dissociation constant, and the reciprocal of Ka
what are the units of Kd
M
when [L]=Kd, how many of the ligand binding sites are filled
half
how much ligand is bound as [L] falls below Kd
less ligand is bound
how much ligand is bound as [L] rises above Kd
more ligand is bound
a low Kd value corresponds to a (higher/lower) affinity of the ligand for the protein
higher
since the ligand of myoglobin is O2 (gas), what adjustments do we make
we measure the partial pressure of O2 (pO2) instead of concentration
what does P50 mean
the partial pressure of O2 at which 50% of ligand binding sites are occupied
using partial pressure of O2, what is the equation for Y
Y= pO2/pO2+P50
does myoglobin have a low or high affinity for O2
VERY high, due to low Kd value
T or F: the binding of a ligand to any protein is affected by the protein structure and is often accompanied by conformational changes in the protein
true
what different molecules can heme bind to
O2, CO, and NO
describe the affinities heme has for CO and O2 when heme is free
CO has a MUCH higher affinity
describe the affinities heme has for CO and O2 when heme is bound to myoglobin
CO still has greater affinity, but the difference is much less significant
which ligand binds to heme at an angle
O2
which ligand binds to perpendicularly
CO
which angle of binding is stronger: straight or angled
straight
will O2 or CO be more stabilized when bound to heme
O2, via the distal His. CO will not be stabilized by anything
what happens if CO outcompetes O2 to bind to heme
it can kill us
