Download Protein Structure and Function - Wk 1-2

Alistair LOs_Protein Structure and Function_Wk 12 Wine Dark Sea
Define primary, secondary and tertiary structure of protein, using haemoglobin as an example.
The primary structure
The primary structure of a protein is the linear sequence of amino acids. Individual amino acids are joined by
peptide bonds, which are amide linkages between the -carboxyl group of one amino acid and the -amino
group of another. The free amino end of the protein is the N-terminus while the free carboxyl end is the Cterminus.
Secondary Structure
The secondary structure of a protein refers to the local structure of the polypeptide chain. This structure is
determined by hydrogen bond interactions between the carbonyl oxygen group of one peptide bond and
the amide hydrogen of another nearby peptide bond. There are two types of secondary structure, the αhelix and the β-pleated sheet.
The α-helix is a rod-like structure with the peptide chain tightly coiled and the side chains of amino
acid residues extending outward from the axis of the spiral. Each amide carbonyl group is hydrogen-bonded
to the amide hydrogen of a peptide bond that is four residues away along the same chain. There are on
average 3.6 amino acid residues per turn of the helix, and the helix winds in a right-handed (clockwise)
manner in almost all natural proteins. (The amino acid units on a peptide chain are referred to as amino acid
residues. A peptide chain consisting of three amino acid residues is called a tripeptide,)
-pleated sheets are formed by two peptide chains running side by side. These are planar/flat. All
components of the peptide bond in -pleated sheets and are involved in H-bonding. The individual peptide
chains can either run parallel or anti-parallel to one another, the parallel sheet being more stable. -pleated
sheets also occur in both fibrous and globular proteins.
Tertiary structure
The three-dimensional, folded and biologically active conformation of a protein is referred to as its tertiary
structure. It consists of complex coiling and folding and gives the protein its final 3D shape. Hydrophobic
groups are buried in the centre and interact with each other via hydrophobic interactions away from water.
Hydrophilic residues generally accumulate on the proteins surface and interact with each other and water.
The tertiary structure is stabilised by amino acid interactions including hydrogen bonds (primary force),
covalent disulfide bonds between adjacent cysteine residues (these bonds create permanent loops or coils),
ionic interactions and hydrophobic interactions.
• Hydrophobic groups are buried in the centre and interact with each other via hydrophobic interactions
away from water.
• Hydrophilic residues generally accumulate on the proteins surface and interact with each other and water
Explain how protein structure relates to function, using the cytochromes, myoglobin and haemoglobin as
examples
Don’t forget that: Alter the sequence alters the structure which alters the function
General Protein Structure
Fibrous Protein - linear and have repeated primary sequences and secondary structure
– eg. keratin, collagen
Globular Protein - globular, roughly spherical in shape, they can have regions of -helix,
-sheet and random coil – eg. enzymes and receptors
Cytochromes, found in the mitochondrion and endoplasmic reticulum, are proteins that contain heme
groups which transport electrons. They are not involved in oxygen transport. In hemoglobin and myoglobin,
heme must remain in the ferrous (Fe2+) state; in cytochromes, the heme iron is reversibly reduced and
oxidized between the Fe2+ and Fe3+ states as electrons are shuttled from one protein to another. The Fe ion
is located right in the middle of the four 5-membered rings (of the haemoglobin) with the pyrrole nitrogen
from each ring adjacent to it. The Protein component contains alpha-helices.
Myoglobin: Can only bind one molecule of O2 as it only has one heme group. Myoglobin consists of 8 helices
arranged into one globular protein. It reversibly binds an oxygen molecule so as to transport it by diffusion
from the edge of a muscle cell to a mitochondria where the oxygen will be used. Myoglobin has a higher
affinity for O2 than haemoglobin. So the dissociation curve is shifted to left of Hb. That is at the same partial
pressure of O2 (pO2) the percent saturation of myoglobin will be higher than that for haemoglobin.
Myoglobin has a hyperbolic dissociation curve owing to the fact it binds only one O2. As pO2 decreases the
reaction is shifted left to release O2 and if pO2 is increases the reaction is shifted right and O2 stays bound.
Note that the high affinity of myoglobin makes it ideal for meeting the O2 demand in the low pO2 conditions
found in active muscle.
Haemoglobin The solubility of oxygen in blood plasma is very low so it requires a transporter. Haemoglobin
transports O2 and CO2 throughout the body within the RBCs (~70%) and is a tetramer of 2 alpha and 2 beta
subunits, 22 in adults.
It can bind four O2 molecules as it contains 4 haemoglobin subunits each with 1 heme group. Each heme
group bind O2 through an Fe2+ ion. The iron atom remains in Fe (II) ferrous oxidation state whether or not
the heme is oxygenated (Fe II can be oxidised to Fe III to form methaemoglobin which does not bind O2 –
erythrocytes contain methaemoglobin reductase to convert the methaemoglobin that occurs)
The heme in each subunit is ligated to one particular His (proximal His). The second His near the Fe ion on
the opposite side of the heme (distal His) but not close enough to be a ligand of the Fe2+ ion. Oxygen binds
Fe2+ to form oxyHb. There is a conformational change when deoxyHb (T state)  oxyHb (R state)
Once one O2 is bound it is easier for subsequent O2 to bind. ie. There is an increase in affinity for O2. The
positive cooperativity of O2 binding to Hb arises from the effect of the ligand-binding state of one heme on
the ligand-binding affinity of another. Therefore the O2-Hb dissociation curve is sigmoidal.
Outline the differences in sub-unit structure and function of foetal and adult haemoglobin in terms of
affinity for oxygen and 2,3 BPG
Sub-unit structure
Function & Affinity for O2
Affinity for 2,3bisphosphoglycerate
FOETAL (HbF)
α2 2
(but once born, β replaces )
GREATER (shifts O2 binding curve LEFT)
Affinity needs to be higher than
mother’s otherwise won’t be able to
‘steal’ O2 from mother. Mother’s
circulation has PO2 of about 50mmHg
\foetus has to have greater affinity to
take O2 from mother’s Hb.
LESSER affinity for 2,3-BPG \can have
greater affinity for O2! Lesser affinity
ADULT (HbA)
α2 β2
(can also get HbA2 which is α2 δ2 if not
enough β present eg. thalassaemia)
LESSER compared to HbF but balances O2
affinity with ability to RELEASE O2 to
tissues and LOAD O2 at the lungs.
GREATER affinity for 2,3-BPG. This isn’t a
bad thing though because need BPG to
(BPG)
because  globin chain has Ser(-ve
charge) instead of β globin chain which
has His (+ve charge) \less likely for (–)ve
BPG to bind.
LESSER Hb’s affinity for O2 so that makes
it easier to RELEASE O2. That’s why in
high altitude, the body
GREATER BPG so that can release more
O2 to tissues.
2,3-bisphosphoglycerate (2,3-BPG) reduces haemoglobin’s affinity for O2
2,3-Bisphosphoglycerate (2,3-BPG) is an important by-product of glycolysis in the RBC. 2,3-BPG is a negative
allosteric effector of the O2 affinity of Hb. It decreases the O2 affinity of deoxyhemoglobin, promoting the
release of O2 in peripheral tissue.
BPG binds tightly to deoxy Hb in a 1:1 ratio, but only weakly to oxy Hb so the presence of BPG decreases
haemoglobin’s oxygen affinity by keeping it in the deoxy conformation.
In arterial blood, where pO2 is 100, Hb is 95% saturated, but in venous blood, where pO2 is 30, it is 55%
saturated
 In passing through capillaries, Hb unloads 40% of its oxygen
 In the absence of BPG the amount released is decreased, as Hb has a high oxygen affinity and will
bind tightly to it
 In the presence of BPG, more oxygen is released as it decreases the Hb’s affinity for O2.
 CO2 and BPG independently modulate Hbs O2 affinity
In anemia and high altitudes, more BPG is produced, which cannot exit the erythrocyte membrane, and thus
causes a decrease in O2 binding affinity, meaning more is unloaded at the capillaries
Foetal haemoglobin (HbF) has a higher affinity for O2 than adult haemoglobin (HbA), due to HbF’s lower 2,3bisphosphoglycreate affinity.
This is due to the fact the -globin chains of HbF lack some positively charged amino acid residues found in the globin chains that are responsible for binding 2,3-bisphosphoglycerate.
Note that HbA and HbF have similar O2 affinities if stripped of 2,3-bisphosphoglycerate but the amino acid
differences in the -globin chains of HbF means that in the presence of 2,3-bisphosphoglycerate HbA is at an O2
binding disadvantage.
The greater O2 affinity of HbF allows more efficient transport of O2 across the placenta to the RBCs of the foetus
as the mother’s blood is less saturated with O2 as it has already passed through part of he circulation as opposed
to adults who easily pick up O2 from the lungs.
Unloading of O2 to tissues is decreased (harder for O2 to be released from the Hb), and causes a leftward shift in
dissociation curve compared with HbA.