Protein Function
Structure will determine the function
of the protein
Key ideas and terms
• protein can bind a ligand in the binding site
• For an enzyme, the ligand is a substrate and
they bind in what is called the active site
• ligand has to be the correct shape
• ligand has to have the complementary
charges and hydrophobicity or
hydrophilicity
Lock and Key Hypothesis
• Protein and ligand have
complementary shapes.
• Interactions must also be
complementary
– If enzyme charge is
negative, substrate must be
positive
– If pocket is nonpolar, ligand
must be nonpolar
• Antibodies
Induced Fit
• Induced Fit: when the
protein and ligand
bind, the protein may
change conformation
to allow for tighter
binding
• Frequently, both the
ligand and the protein
change conformation
Examples: O2 binding proteins:
myoglobin and hemoglobin
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oxygen is not water soluble yet needs to be transported
diffusion is not effective
myoglobin is found primarily in muscle tissue
Hemoglobin is in the blood
Both proteins contain heme
Heme Group
consists of a Fe2+ and a protoporphyrin ring to help stabilize
the iron(II) ion
Heme Group
the iron must be a 2+ to bind oxygen. The heme group is
buried deep within the protein so that the iron is not
oxidized to 3+
• there must be flexibility in the protein to allow for oxygen
to attach and then let go
• Iron has 6 coordination sites.
– Four of them used by porphyrin. Unshared pairs on nitrogen
complex to iron
– Fifth and 6th for oxygen and protein
• Heme is planar
Myoglobin
• has heme group
• eight alpha helical segments
• dense hydrophobic core
– all but two polar groups on
outside
– room for only 4 water
molecules
• flat heme in pocket
• iron coordinated to poryphorin
and H
• As well as the heme
Myoglobin Binding Curve
• Hyperbolic binding
curve
• Relatively insensitive
to small changes in
oxygen concentration
Myoglobin
• The P50 (oxygen partial pressure required for half
saturation) for myoglobin is very low
• Myoglobin has a high affinity for oxygen-an important
characteristic for a protein that must extract oxygen from
the small amounts present in blood.
• At the oxygen concentration existing in the capillaries, the
myoglobin in adjacent tissues is nearly saturated.
• When cells are metabolically active, their internal PO2 falls
to levels where myoglobin will lose (deliver) its oxygen.
Hemoglobin
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Quaternary structure : 4
subunits
Each subunits is like
myoglobin
Each subunit has heme
group
2 alpha chains; 2 beta
chains
Few contacts between
alpha and betas, more
between alphas and betas
Hemoglobin
• Exists in two states
– R state (high affinity for O2)
– Where would this state be favored?
• In the lungs
– T state (low affinity for O2) (deoxyhemoglobin)
– Where would this state be favored?
• In the tissue
– Sensitive to pressure changes
– On oxygenation, one pair of subunits shifts with
respect to the other by a rotation of 15 degrees.
Hemoglobin
• deoxy hemoglobin (T)
oxy hemoglobin (R)
Oxygen Binding to Heme in
Hemoglobin
• Fe is coordinated to a histidine
in helix 8 of the Hb molecule
• In deoxy form, porphyrin is
puckered and Fe is out of the
plane of the heme
• When oxygen binds the Fe (at
other coordination site) the Fe
is pulled into the plane of the
heme
• This pulls on the histidine,
which pulls on the helix,
changing the shape of the
molecule.
His F8
0.6 A
Fe2+
O2
Hemoglobin
• Conformational changes in hemoglobin alter its binding
ability
• Binding of oxygen in one subunit causes conformational
changes in the next subunit
• This is called cooperative binding
• This can happen because it is composed of 4 independent
subunits
• produces a different binding curve that is sigmoidal
• The modulation of the affinity of a site for a ligand by
ligand binding at another site is called Allostery.
Hemoglobin Binding Curve
Bohr Effect
• Hemoglobin's affinity for oxygen is decreased in the
presence of carbon dioxide and at lower pH.
• Carbon dioxide reacts with water to give bicarbonate,
carbonic acid free protons via the reaction:
CO2 + H2O ---> H2CO3 ---> H+ + HCO3-
• Protons bind at various places along the protein and carbon
dioxide binds at the alpha-amino group forming carbamate.
• This causes a conformational change in the protein and
facilitates the release of oxygen.
Bohr Effect
• Blood with high carbon dioxide levels is also lower in pH
(more acidic). (recall the equilibrium)
• Conversely, when the carbon dioxide levels in the blood
decrease (i.e. around the lungs), carbon dioxide is released,
increasing the oxygen affinity of the protein.
Bohr Effect Summary
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High CO2 in tissues
Higher H+
Lower pH
Affinity for O2
decreases
• O2 released to tissues
• T state favored
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Low CO2 in lungs
Lower H+
Higher pH
Affinity for O2
increases
• O2 binds hemoglobin
• R state favored
Hemoglobin and CO poisoning
• Other ligands can compete with oxygen for binding to
hemoglobin
• The binding of oxygen is affected by molecules such as
carbon monoxide (CO) (For example from tobacco
smoking, cars and furnaces). CO competes with oxygen at
the heme binding site.
• Hemoglobin binding affinity for CO is 200 times greater
than its affinity for oxygen, meaning that small amount of
CO can dramatically reduces hemoglobin’s ability to
transport oxygen.
• Hemoglobin also has competitive binding affinity for
Nitrogen Dioxide and Hydrogen sulfide .
Oxygen and Carbon Monoxide
• Oxygen and carbon
monoxide same size
and shape.
• Carbon monoxide,
however has formal
charge
• Sticks to Fe better
• Blocks oxygen
binding
Fe
Fe
2+
2+
O
O
+
C
O
Hemoglobin and 2,3 DPG
• In people acclimated to high
altitudes, the concentration of
2,3-diphosphoglycerate (2,3DPG) in the blood is increased,
which allows these individuals
to deliver a larger amount of
oxygen to tissues under
conditions of lower oxygen
tension.
• This phenomenon, where
molecule Y affects the binding
of molecule X to a transport
molecule Z, is called a
heterotropic allosteric effect.
O
O
O
C
H C
O
CH2OH
P O
O
O
P O
O
Sickle Cell Anemia
• Sickle cell disease is caused by an abnormal
adult hemoglobin, called hemoglobin S.
People with sickle cell disease make
hemoglobin S instead of hemoglobin A.
• Single amino acid substitution
– glutamate changed to valine
• To show condition, have to have mutation in
both genes (Homozygous)
• http://www.scinfo.org/sicklept.htm
Sickle vs normal hemoglobin
• first 9 amino acids of normal hemoglobin
beta chain
vhltpeeks
• first 9 amino acids of sickle hemoglobin
beta chain
vhltpveks
Notice the single amino acid change?
Sickle Cell Anemia
• Position 6 is on outside of molecule
• Glutamate is polar
• Valine substitution causes “sticky spot” on
outside of hemoglobin
• Causes hemoglobin molecules to stick
together
• Forms long chains which cause red blood
cell to sickle