Transcript Ch. 7 Protein Function and Evolution Myoglobin and Hemoglobin • Both are essential for oxygen need • Myoglobin stores O2 in the muscle •

Ch. 7 Protein Function and
Evolution
Myoglobin and Hemoglobin
• Both are essential for oxygen need
• Myoglobin stores O2 in the muscle
• Hemoglobin transports O2 to tissues and CO2
and H+ back to lungs
• The 2o structure of hemoglobin resembles
myoglobin but the 4o structure allows for
interactions that are central to its function.
Structure
• Figure 7.3 page 214
– Notice Hemoglobin appears to be constructed of 4
Myoglobin strands
• Figure 7.4 page 215
• Heme is composed of 4 pyrrole rings linked by
a-methylene bridges.
• As a whole, the molecule is called Porphin
• Each Porphin binds 1 Ferrous Ion (Fe2+)
Structure, cont.
• Oxidation of the iron to Fe3+ destroys
biological function
Myoglobin
• Oxygen stored is released to prevent oxygen
deprivation
• The oxygen goes to the mitochondria for
synthesis of ATP
• Myoglobin is composed of about 75% alpha
helixes which is unusually high
• As with most globular proteins, outside is
polar and inside is non-polar
His at E7 and F8
• The eight helixes are termed A-H starting at the
amino terminal
• Notice the Histidine residues at E7 and F8, near the
site of Heme (porphin system) Figure 7.5 page 216
• His contains pyrrole like ring as a side chain
• Heme binds to myoglobin with the propanate
groups towards the outside (polar) and the
nonpolar methyl and vinyl groups towards the
inside (see fig 7.4c)
Histidines
• The F8 His actually provides a fifth
coordination to the iron providing an actual
linkage to the protein. (fig 7.5b)
• The other histidine, E7, lies on the opposite
side. (more on this later!)
• Due to the coordination to the F8 His, the iron
lies outside the plane of the heme and
puckers the heme slightly
Heme binding of O2
• When the iron binds O2, the iron moves closer
to the plane, pulling the F8 His with it, thus
slightly altering the other residues near the F8
His.
• When the O2 binds, the preferred orientation
is with the O-Fe-Heme bond at 90o and the
Fe-O-O bond at 121o
Heme binding to CO
• The iron actually binds CO with a similar bond
that is 25,000 times stronger!
• However, the C of the CO is sp hybridized and
so the Fe-C-O bond should be 180o
• This angle is not allowed due to the presence
of the E7 Histidine.
• There is a lone pair of electrons on the
Nitrogen that creates steric and electronic
repulsions
Heme binding to CO
• As a result, CO is forced to bond like O2 and
the C-O-Fe bond is significantly weakened.
• This weakening allows for the great
abundance of O2 to predominately bind.
Myoglobin: Storage vs. Transport
• Myoglobin is better for storage than transport
• The reasoning is seen in the Oxygen binding
curve. (fig 7.6, page 217)
• Notice how the % saturation doesn’t begin to
drop until the PO2 is very low.
• This means that Myoglobin would not release O2
in normal conditions, only in very low levels of O2
Hemoglobin
• The additional properties of hemoglobin that
allow it to effectively transport O2 arise from it
4o structure.
• These are referred to as allosteric properties,
meaning “other space”
• Hemoglobin is tetrameric and contains 2 pairs
of different peptide sub units
• The 1o structure of b,g,d are highly conserved
Comparisons
• Myoglobin and b-subunits have almost
identical 2o and 3o structures
• The a strand is also similar but only contains 7
helices rather than 8
Hemoglobin
• Hemoglobin contains 4 heme groups,
therefore it can bind 4 O2 molecules per 1
hemoglobin
• Recall that the binding of O2 slightly changes
the structure of the heme and connecting
protein
• This slight change allows for the next O2 to
bind easier
• This is called cooperative binding
• Cooperative binding helps hemoglobin both
load and unload O2
• Cooperative binding is only seen in multimeric
proteins.
P50
• P50 is the quantity used to express O2 partial
pressure
• P50 is the partial pressure of O2 that half
saturates a given hemoglobin
• P50 will vary organism to organism but will
always exceed the PO2 in peripheral tissues
Cooperative Binding
• The reason hemoglobin experiences
cooperative binding is the large
conformational changes that hemoglobin
undergoes when O2 is bound
• When O2 in bound, one of the a/b subunits
rotates 15o creating a more complex structure
• The relates to profound changes in the 2o, 3o,
and 4o structure
Conformations
• Hemoglobin with no O2 bound is said to be in
the T (taut) form. (Fig 7.13, page 225)
• Once O2 is bound, the hemoglobin shifts to
the R (relaxed) form.
• This conformational shift is what lowers the
binding energy for the remaining O2 to bind
• Less conformational change is needed.
The Return Trip
• Hemoglobin not only transports O2 from the
lungs to peripheral tissues, but also transports
CO2 and H+ from the peripheral tissues back to
the lungs
• The CO2 is the by-product of respiration in
cells
• The CO2 does not bind to the same sites as O2.
Transport of CO2
• CO2 forms carbamates with terminal amino
groups of the proteins of Hemoglobin
• This binding of CO2 changes the charge at the
N-terminal from + to –
• This favors additional salt bridges holding
hemoglobin together.
Transport of CO2
• Only about 15% of CO2 in transported in this
manner
• Most of the rest is transported as bicarbonate
• Bicarbonate is formed in erthrocytes by the
hydration of CO2 which is catalyzed by carbonic
anhydrase
• Initially, carbonic acid is formed but immediately
deprotonates at the pH of the blood
Acidic Environment
• Hemoglobin will bind one H+ for every 2 O2’s
released
• This plays a major role in buffering capacity of
blood
• The delivery of O2 is enhanced by the acidic
environment of the peripheral tissues due to
the carbamation stabilizing the T form.
In Lungs
• In the lungs the whole process is reversed!
• The reciprocal coupling of H+ and O2 binding is
termed the Bohr effect.
Bohr Effect
• The Bohr Effect is dependent upon the
cooperative interactions between the hemes
of the tetramer
• Therefore, Myoglobin would not show the
Bohr Effect
• So, where do the protons in the Bohr Effect
come from and how do they help enhance the
release of O2?
You had to ask!!!!
Other Factors
• Release of O2 is also enhanced by the
presence of 2,3-biphosphoglycerate (BPG)
• BPG is synthesized in erythrocytes at the low
O2 concentrations at peripheral tissues
• BPG helps stabilize the T form of hemoglobin
• It binds in the central cavity formed by the
four subunits of hemoglobin (Fig 7.18 p 229)
• Only the T form binds BPG
• The space between the H helices of the b
chains that line the cavity sufficiently opens
only in the T form
• BPG forms salt bridges with the positive
charges on the terminal amino groups of both
b chains via NA1 (1) and with Lys EF6(82) and
His H21 (143).
• These salt bridges must be broken to return to
the R state.