Chapter 7: Heme Chemistry Flashcards
1
Q
Heme
A
Ferrous protoporphyrin IX
2
Q
Structure of hemoproteins
A
4 parole rings united by 4 methenyl (methene) bridges.
3
Q
Methyl
A
-CH3
4
Q
Vinyl
A
-CH=CH2
5
Q
Propionate
A
-CH CH2 COOH
6
Q
Hemoproteins
A
- Hemoglobin.
- Myoglobin.
- Cytochromes.
- Catalase.
- Peroxidase.
- NO synthase.
- Tryptophan pyrolyse.
7
Q
Myoglobin and hemoglobin
A
Conjugated proteins formed of heme attached to a basic protein (polypeptide chain).
8
Q
Myoglobin
A
1 heme and 1 polypeptide chain (apomyoglobin).
9
Q
Hemoglobin
A
4 heme and 4 polypeptide chains (globin).
10
Q
How many helical does each polypeptide chain is formed of?
A
7-8 helices.
11
Q
How are the amino acids named?
A
- Each helix is given a letter A, B, C… starting from the N terminal.
- Each amino acid is termed by a letter indicating its helix and a number indicating its position in the helix starting from the N terminal.
- Eg. Histidine F8.
12
Q
Heme site
A
Present in a pocket between helix E and F called heme pocket.
The 2 propionate projects outward on the surface.
13
Q
How is heme attached to a polypeptide chain?
A
- Histidine F8 (proximal histidine).
- Hydrophobic interactions with surrounding non polar amino acids.
14
Q
How many coordinations can Fe+2 of heme form?
A
5 or 6 coordinations:
1. 4 coordinations with the 4 nitrogen’s of the 4 pyrrole rings ( in the porphyrin ring).
2. 5th coordination with proximal histidine ( histidine F8).
3. 6th coordination with O2.
Both 5th and 6th coordination are perpendicular to the plane of the ring.
15
Q
Importance of protein parts: apomyoglobin and globin.
A
- Makes heme soluble.
- Prevents diffusion of the heme from RBC’s to plasma.
- Keeps iron in ferrous state (heme pocket is surrounded by non polar amino acids) and prevents oxidation of heme into hematin and prevent formation of heme oxygen heme complex.
- Decreases affinity of heme to CO from 25,000 to 200.
- Responsible for the sigmoid shape of O2 dissociation curve of hemoglobin.
16
Q
Decreases affinity of heme to CO 25,000 to 200
A
- Heme has a higher affinity to CO than O2.
- CO binds to heme at the same binding site of O2 which will lead to hemoglobin not being able to carry O2.
- The angle between 1st and 2nd oxygen atoms is about 121 degrees.
- The preferred position for CO is:
- Fe
- C
- O
Perpendicular to the plane of the ring. - When O2 binds to heme, the 1st oxygen perpendicular to the plane of the ring and the bond between and 1st and 2nd oxygen is 121 degrees.
- Distal histidine in polypeptide chain produces steric hindrance stabilizing O2 binding and destabilizing CO binding.
- CO is produced in very small amounts in the body from catabolism of Hb.
17
Q
Myoglobin site
A
Muscles (skeletal and cardiac).
- Gives muscles it’s red color.
18
Q
Myoglobin function
A
Storage of O2.
19
Q
Myoglobin affinity to O2
A
Higher affinity to O2 than hemoglobin.
20
Q
Myoglobin structure
A
1 heme and 1 polypeptide chain (apomyoglobin).
21
Q
Apomyoglobin
A
- 153 aa.
- 8 helices (A to H).
- Soluble in water:
Polar amino acids are present outside and non polar amnio acids are present inside (wavy interior) except for Histidine F8 and E7.
22
Q
Hemoglobin site
A
RBC’s
23
Q
Hemoglobin function
A
- Carry O2 from lung to tissues and CO2 from tissues to lung.
- Hb/HbO2 system acts as a buffer in RBC’s.
24
Q
Hemoglobin affinity to O2
A
Lower affinity to O2 than myoglobin.
25
Hemoglobin structure
1. 4 heme and 4 polypeptide chains:
- Hb A: 98%.
- Hb A2: 2%.
26
Hb A
Major adult hemoglobin.
27
Hb A globin
Formed of 4 polypeptide chains (2a and 2B) arranged in 2 dimers (aB)1 and (aB)2.
Each dimer is formed of 1a and 1B held together by hydrophobic interactions.
28
The 2 dimers are less tightly held together by?
1. Hydrogen bond.
2. Ionic interactions (salt bridges).
29
Globin structure number
Tetrameric.
30
How many helices is the alpha chain of hemoglobin formed of?
141aa arranged in 7 helices.
31
How many helices is the beta chain of hemoglobin formed of?
146 aa arranged in 8 helices. (Similar to apomyoglobin).
32
How many forms of hemoglobin are there?
2 forms: R form and T Form.
33
How does the rotation of dimers occur?
As the 2 dimers are less tightly bound together, this allows the rotation of 1 dimer 15 degrees relative to the other by oxygenation.
34
Ability of hemoglobin to reversible bind with O2 is affected by?
1. O2.
2. PH of the environment (Bohr effect).
3. CO2.
4. Availability of 2,3 biphosphoglycerate (2,3 BPG).
5. CO.
All these factors are called Allosteric effectors. Binding of these effectors at one site on Hb affects binding of O2 to heme groups at other locations of the molecule.
35
Oxygen binding site
O2 binds to Hb and lies in O2 binding site (O2 pocket).
36
Oxygen is coordinated to?
Fe+2 (6th coordination position).
37
What stabilizes the binding of O2 to Fe+2?
Histidine E7 (distal histidine).
38
Oxygenation
1. A cooperative process: oxygenation helps more oxygenation.
2. Step 4 is easier than step 3, step 3 is easier than step 2 and so on.
3. The affinity of Hb to the last O2 is 300 times greater than the 1st O2.
39
What is the cause of the sigmoidal shape of O2?
Cooperative binding of O2 to Hb is the cause of the O2 dissociation curve of Hb.
40
Why is the cooperative binding of O2 the cause of the sigmoidal shape?
Binding of O2 to 1 of the polypeptide chains will lead to rupture of 2 salt bridges which increases the affinity of other subunits to O2. So, when all subunits bind to O2, it will lead to rupture of 8 salt bridges and Hb is converted from T form to R form.
41
T form oxygenation
Deoxygenated
42
R form oxygenation
Oxygenated
43
T form ionic bonds
More ionic bonds.
44
R form ionic bonds
Less ionic bonds
45
T form mobility
Less mobile
46
R form mobility
Rotates 15 degrees
47
T form affinity to O2
Lower affinity to O2
48
R form affinity to O2
High affinity to O2
49
T form is stabilized by?
1. Deoxygenation.
2. H+ (protonation).
3. CO2.
4. 2,3 BPG.
50
R form is stabilized by?
1. Oxygenation.
2. Deprotonation.
3. Removal of CO2 and 2,3 BPG.
51
T form name
Tense or Taut
52
R form name
Relaxed
53
Changes accompanied with the transition of T form to R form: absence of O2
1. Porphyrin ring is slightly curved upwards.
2. Fe+2 lies above the plane of the ring by 0.04 nm.
54
Binding of O2 causes:
1. Porphyrin ring is flattened.
2. Oxygenation of Hb is associated with rupture of 8 salt bridges.
3. The 2 dimers rotate upon each other by 15 degrees.
4. Pulling of Fe2+ pulls proximal histidine pulls helix F, which release 2 protons from imidazole ring of histidine 146 of B chain and N terminal amino groups.
5. The liberated hydrogen ions bind with bicarbonate and forms carbonic acid that is broken by carbonic anyhydrase to CO2 (lost by the lungs) and H2O.
55
Most of the CO2 delivered from tissues to the blood is converted to?
85% of CO2 is converted to H2CO3.
56
How is O2 released in the Bohr effect?
1. Most (85%) of CO2 delivered from tissues to the blood is converted to H2CO3.
2. In RBC’s, H2CO3 liberates hydrogen ions and HCO3.
3. H+ binds to the imidazole ring of histidine 146 and N terminal amino group, stabilizing T form so the affinity to O2 is decreased helping the release of O2.
The reverse occurs in the lungs.
57
2,3 biphophoglycerate
A negative charged molecule that binds to the positive charges pocket in the center of 4 Hb subunits.
58
Binding of 2,3 BPG results in?
Formation of more ionic bonds which favors T form.
59
2,3 BPG at tissues
1. Low O2 tension.
2. Binding of 2,3 BPG stabilizes T form, which releases O2. It results in the oxygenation of tissues.
60
2,3 BPG at lungs
1. High O2 tension.
2. Binding of O2 releases 2,3 BPG which stabilizes R form.
3. 2,3 BPG causes shift of oxygen dissociation curve to the right.
61
Carbamino Hb
1. Most (85%) of CO2 is transported to lungs as bicarbonate ion.
2. 15% of CO2 formed at tissues is carried in the form of carbamino Hb on the N terminal a-amino group with release of H+.
CO2 + Hb - NH2 = Hb- NHCOO- +H+
62
CO bind to Hb but reversibly… why?
1. Carbon monoxide binds at O2 binding site.
2. Affinity of Hb to CO is 200 times a that of O2.
3. When CO binds to 1 or more of the 4 heme sites, Hb shifts to the relaxed R form, so the remaining heme site bind O2 with high affinity which makes Hb unable to release O2.
4. So CO is very toxic.
5. CO poisoning is treated by 100% oxygen therapy which helps dissociation of CO from Hb.
63
Minor hemoglobins
1. Embryonic hemoglobin (Hb gower).
2. Fetal hemoglobin (HbF).
3. HbA2.
64
Hb Gower structure
1. Gower 1: ( Zeta 2, epsilon 2).
2. Gower 2: (alpha 2, epsilon 2).
65
Hb Synthesis
During the 1st 3 months of pregnancy.
66
Hb Gower characters
1st hemoglobin formed during embryonic life.
67
Hb Gower characters
1st hemoglobin formed during embryonic life.
68
HbF structure
Alpha2, Gamma 2
69
HbF synthesis
After 3 months of pregnancy.
70
HbF affinity to O2
Higher affinity to O2 than maternal Hb, so it allows fetal Hb to take O2 from maternal blood. For example: HbF shift oxygen dissociation curve to the left.
71
HbF electrophoretic mobility
Slower in electrophoresis than HbA.
72
HbA structure
Alpha 2, Beta 2
73
HbF structure
Alpha 2, Gamma 2
74
HbA time
From 6 months of pregnancy till end of life.
75
HbF time
From 3 to 6 months of pregnancy.
76
Electrophoresis
Faster in electrophoresis.
77
Electrophoresis
Slower in electrophoresis.
78
HbA Affinity to O2
Lower
79
HbF affinity to O2
Higher
80
Affinity to 2,3 BPG
Higher
81
HbF affinity to 2,3 BPG
Lower due to replacement of histidine by serine in Gamma chain.
82
HbA1c binding site
1. Glucose binds to the N terminal valine of the B chain.
2. Binding of glucose to Hb occurs spontaneously and non enzymatic.
83
What does the rate of binding of HbA1c depend on?
It depends on the level of blood glucose.
It reflects the level of blood glucose over the last 60 days (6-8 weeks).
84
What is the range of blood HbA1c in a normal person and controlled diabetics?
X<6.5%
85
Range of HbA1c of a diabetic person.
X>6.5%
86
Range of HbA1c in an uncontrolled diabetes mellitus.
X>8%
87
Glyclated Hb
HbA1c
88
What is HbA1c used for?
It is used for diagnoses and follow up of diabetes mellitus.
89
Myoglobin site
Muscle
90
Hemoglobin site
RBC’s
91
Myoglobin function
Storage of O2
92
Hemoglobin function
1. Carry O2 from lung to tissues and CO2 from tissues to lung.
2. Buffer function.
93
Myoglobin structure
1 heme and 1 polypeptide chain.
94
Hemoglobin structure
4 heme and 4 polypeptide chains.
95
Myoglobin number of O2 molecules
1
96
Hemoglobin number of O2 molecules
4
97
Myoglobin affinity to O2
High affinity
98
Hemoglobin affinity to O2
Lower affinity
99
Types of myoglobin
1
100
Types of hemoglobin
1. HbA
2. HbA2
3. HbF
101
Organization of the globin gene families
Genes encoding for a gene family and B gene family are present on separate genes on 2 separate chromosomes.
102
a gene gene family site
Chromosome 16
103
A gene family contains
1. Zeta gene: during 1st 3 months.
2. 2 alpha (alfa) genes (a1 and a2): encodes for a chain.
104
B gene family site
Chromosome 11
105
B gene family contains
1. Epsilon gene: 1st 3 months of pregnancy.
2. 2 gamma genes: HbF.
- Ay: aa No. 136 alanine.
- Gy: aa No. 136 glycine.
3. Delta gene: HbA2.
4. Beta gene: HbA.
106
Hemoglobinopathies
Family of disorders resulting from:
1. Production of abnormal hemoglobin.
2. Decreased production of normal hemoglobin (thalassemia).
107
Production of abnormal hemoglobin
1. Sickle cell anemia (HbS disease).
2. HbC disease.
3. HbM disease (methemoglobinemia).
108
Decreased production of normal hemoglobin (thalassemia).
1. a thalassemia: defect in the production of a chain.
2. B thalassemia: defect in production of the B chain.
109
Sickle cell anemia
Genetic disease due to single nucleotide substitution (point mutation).
110
What happens in sickle cell anemia?
1. Amino acid no.6 in B chain glutamate is replaced by valine.
2. Hydrophobic nature of valine produces protein with marked tendency to form insoluble aggregates due to presence of sticky patches in the B chain and receptor for sticky patches in the a chain.
3. Polymerization is more in cases of deoxyhemoglobin due to presence of sticky patches on B subunits and their receptors on (a) subunits.
111
How many genes are their for the B chain?
2 genes for the B chain (1 in each chromosome 11):
1. Homozygous for sickle cell anemia: HbS.
2. Heterozygous for sickle cell anemia: HbA and HbS.
112
Effects of sickle cell anemia
1. RBC’s acquire sickle shape.
2. Decrease in life span of RBC’s: due to increased removal of sickle RBC’s by spleen at a faster rate.
3. Frequent infarctions: aggregated RBC’s block blood vessels which leads to localized anoxia, severe pain, and even death.
4. Individual becomes resistant to malaria due to decreased life span of RBC’s: parasites can’t complete its life cycle.
113
HbC
1. Genetic disease.
2. Amino acid number 6 in B chain (glutamate) is replaced by lysine.
3. RBC’s crystallize which decreases its life span and leads to mild anemia.
114
Hereditary HbM
1. Genetic disease.
2. Proximal or distal histidine is replaced by tyrosine.
3. Fe2+ in converted to Fe3+.
4. Heme is converted to hematin.
5. Hb in converted to met Hb which can’t carry O2.
115
Acquired HbM
HbM may result normally in vivo from oxidation of Hb by:
1. Free radicals: H2O2.
2. Drugs:
- Sulfonamides.
- Nitrites.
- Nitrates.
3. Decreased activity of NADH met Hb reductase: normally reverses the oxidation of Hb.
116
Thalassemia
Hereditary hemolytic disease due to gene mutation or deletions.
117
How many gene are present in a genes?
4 a genes, 2 on each chromosome 16.
118
a chain: defect in 1 gene
1. Carrier for a thalassemia.
2. Symptoms: completely normal (silent carrier).
119
a chain: defect in 2 genes
1. a thalassemia trait.
2. Symptoms: mild anemia.
120
a chain: defect in 3 genes
1. a thalassemia major.
2. Symptoms: severe anemia.
121
a chain: defect in 4 genes
1. Homozygous for a thalassemia.
2. Effects:
- Dies intrauterine (hydrops fetalis).
- Formation of abnormal Hb which have high affinity to O2:
* Hb Bart (y4).
* Hb H (B4).
122
How many B globin genes are present in each chromosome?
2 B globin genes. 1 on each chromosome 11.
123
B chain: defect in 1 gene
1. B thalassemia minor (B thalassemia trait).
2. Symptoms: mild anemia.
124
B chain: defect in 2 genes
1. B thalassemia major.
2. Symptoms: severe anemia layer on after birth.
3. Treatment: blood transfusion.
4. Prognosis: death before adulthood.
125
Why does the patient with B thalassemia major appear normal after birth?
Due to presence of HbF (a2y2).
126
Why are there high levels of HbF and HbA2 in patient with B thalassemia?
To compensate the absence of HbA.
127
Embryonic Hb
1. During 1st 3 months of pregnancy.
2. Gower 1: (gamma 2, epsilon 2).
2. Gower 2: (alpha 2, epsilon 2).
128
Fetal Hb (HbF)
1. (Alpha 2, gamma 2) after the 1st 3 months of pregnancy.
2. 80% of total Hb at birth.
129
Adult Hb (HbA)
1. (Alpha 2, Beta 2) major adult Hb.
2. 98% of adult Hb.
130
HbA2
1. (Alpha 2, Delta 2) minor Hb adult.
2. 2% of adult Hb.
131
HbS
6th aa B chain (glutamate) is replaced by valine.
132
HbC
6th aa B chain (glutamate) is replaced by lysine.
133
HbM
Proximal or distal histidine in a or B chain is replaced by tyrosine.
134
Hb Bart
(Y4) in individuals homozygous for a thalassemia.
135
HbH
(B4) in individuals homozygous for a thalassemia.
