Biochemistry And Genetics Of RBCs Flashcards
Structure of RBC overview
Lack of nucleus and membrane bound granules
Shaped in a biconcave disc to maximize the cell surface for oxygen exchange
Short life span (120 days on average) and very fast production (10^12 on average daily)
Hereditary elliptocytosis
RBC assumes an elliptic shape, usually caused by abnormalities in spectrin protein (also possible in band 4.1 and glycoproteins C proteins, but this is less common)
Hereditary spherocytosis
Caused by deficiency and/or abnormalities in spectrin protein (can also becaused by ankyrin, band 4.1, 4.2 proteins but is less common)
Symptoms:
- presence of spherocytes w/ really short lifespan
- hemolytic anemia and splenomegaly
Affects people of Northern European descent usually.
RBC metabolism
Glycolysis:
- to generate ATP
- production of 2,3 BPG
- Reduces Iron
PPP/HMP:
- generates NADPH which is used to fuel G3P and F6P production
- NADPH is used to reduce glutathione which is used to protect RBCs from oxidizing damage
RBC energy metabolism
Only uses anaerobic glycolysis
(does not have mitochondria)
NAD+ is restored via lactate production
Pyruvate kinase deficiency
Results in hemolytic anemia
- 2nd most common deficiency after G6PD
RBCs cant produce lactate so it cant regenerate its NAD+ which is required to generate ATP
- specifically cant generate G3P -> 1,3 BPG
Symptoms:
- fatigue, pale skin, shortness of breath, jaundice, increased gall stone appearance
- NO HEINZ BODIES*
Difference between Pyruvate kinase and G6PD deficiencies
G6PD deficiency produces Heinz bodies (are precipitated hemoglobin)
Pyruvate kinase deficiency does not produce these
What is the most abundant organophosphate in RBCs?
2,3 BPG
Is an allosteric regulator of oxygen binding to hemoglobin and is generated via mutase and 1,3 BPG
- presence increases efficiency of blood cells
Binds to the Beta chains of hemoglobin taut structures.
Is rapidly degraded in blood stored for transfusions and is increased in people that live in high altitudes
How is 2,3 BPG generated?
From 1,3 BPG via mutate enzyme
can enter the Rapoport-Luebering shunt to generate 3 phosphoglycerate via phosphatase if levels are low
PPP (or HMP shunt) steps
Rate limiting step= G6PD
Oxidative reactions are 3 irreversible steps
-purpose is to generate NADPH to reduce glutathione
- nonoxidative reactions are reversible and use two enzymes
- transaldolase
- transketolase
- purpose is to generate nucleotides for precursors
Transketolase activity
Requires TPP (thiamine / Vit. B1) and its activity in RBCs can be directly measured
- low activity of Transketolase = thiamine deficiency
G6PD deficiency
Causes episodic hemolytic anemia induced via oxidative stress within the cell since glutathione cant be reduced
- contains Heinz bodies in affect people
- very common single gene disorder that is x-linked (mainly affects males)
Role of NADPH
Electron donor for biosynthesis of
- Fatty acids
- cholesterol
- steroid
Electron donor for the neutralization of ROS
- hydrogen peroxide
- superoxide
- hydroxyl radicals
Helps reduce Cytochrome P450
Aids in destructions of pathogens by macrophages and neutrophils
Substrate for nitric oxide synthesis
What is the primary role of NADPH in RBCs?
Reducing glutathione (GSH)
- only source of NADPH in RBCs is via PPP and through the oxidation of GSSG from Glutathione reductase
- this is done to make GSH able to fight/ capture free radicals and prevent damage
Heme structure
A Porphyrin cyclic molecule containing 4 pyrrole rings joined via methenyl bridges and a metal ion in the middle
- iron group is ferrous (Fe2) to allow reversible binding to oxygen
Heme biosynthesis and degradation
Most of the heme (85%) is synthesized in bone marrow and requires the presence of iron to do so.
Heme is degraded via mononuclear phagocyte system (MPS) in the spleen, liver and bone marrow
Hemoglobin structure
Two diners composed each of identical heme chains
- strong hydrophobic bonds form between heme chains of each dimer to form stable diners
- each dimer weakly binds to the other via ionic in the deoxygenated state (Taut structure)
- the bonds between each dimer are broken when bound to oxygen (relaxed structure)
- increasing relaxed structure of the hemeglobin as it binds oxygen molecules causes increased affinity for oxygen until all 4 heme groups (positive cooperatively)
Hemoglobin Oxygen dissociation curve
Follows a positive cooperatively model. (The more oxygen bound to heme groups, the high the affinity to oxygen for the non bound heme groups)
- this is caused because hemoglobin becomes relaxed when it binds to oxygen, allowing more open sites for oxygen binding
Oxygen dissociative curve is steepest at the Po2 inside the tissues and at its highest value in the lungs
- sigmoidal curve
Hepcidin
Presence down-regulates transports of iron (tranferrin protein and its receptors on cells) in the GI system and prevents its absorption in the diet.
Hepcidin is turned off when erythropoiesis is needed
primary molecule used in iron metabolism
Hemoglobin vs myoglobin in oxygen dissociation curve
Myoglobin saturates much quicker than hemoglobin due to less total heme groups
- myoglobin P50 = 1mmHg
- hemoglobin P50 = 26 mmHg
P50
The amount of Po2 required to saturate 50% of the hemoglobin/myoglobin
- increases in P50 results in decreased affinity (shift to the right of the curve)
- decreases in P50 results in increased affinity (shift to the left)
Allosteric effectors of hemoglobin affinity to Oxygen
PO2 (increased levels: higher affinity)
PCO2 (increased levels: lower affinity)
PH changes (Lower pH: lower affinity/ Higher pH: high affinity)
2,3-BPG (increased levels: lower affinity)
Bohr effect
Decreases in pH results in decreased oxygen affinity (shift to the right) for hemoglobin since hemoglobin will bind H+ ions (protonates the heme groups) that are produced via bicarbonate
- this causes stabilizing (taut) affects on the deoxygenated hemoglobin, reducing the overall binding sites
PH in lungs is higher than tissues, so oxygen affinity is higher in lungs than tissues
2,3 BPG and hemoglobin affinity
2,3 BPG is an allosteric regulator of O2 and ONLY binds to deoxy-Hb; NOT to oxy-Hb
2,3 BPG binding promotes taut structure and release of oxygen from heme groups, produces a chronic hypoxia effect, but also allows RBCs to release oxygen to the target tissue effectively
Causes right shift in the oxygen saturation curve
Oxygen saturation mechanisms
A measure of the number of occupied O2-binding on a hemoglobin molecule
- independent on the amount of hemoglobin present*
- increases in hemoglobin levels only increase the total amount of O2 content possibly bound
Carbon monoxide saturation mechanisms
Hemoglobin affinity for carbon monoxide is 220x greater than oxygen
- binds tightly (but reversible) to hemoglobin at the same site as oxygen
Presence of CO both reduces o2 carrying capacity and decreases unloading of oxygen from hemoglobin
-It’s binding causes heme sites to very tightly bind to oxygen, decreasing releases as long as CO is present
Cuts the curve peak in half and results in a slight shift towards the left
CO poisoning produces bright cherry red skin since the hemoglobin bound to carbon monoxide turns bright cherry red
Types of hemoglobin in adults
HbA (normal): 90%
HbA2 (2 alpha chains, 2 delta chains): 2-3%
HbF (2 alpha chains, 2 gamma chains): <2%
HbA1c (heme bound to glucose) 4-6%
What is special about HbA1c?
Hemoglobin bound to glucose, high levels (>5.7) suggest prediabetes or diabetes.
Prediabetic levels = 5.7 - 6.4%
Diabetic levels = >6.5%
Expression of hemoglobin genes
Both alpha and beta genes:
- alpha gene only produces alpha chains (zeta only when 0-6 weeks of prenatal age)
- begins at 2 weeks of prenatal age through life*
-beta gene produces gamma, delta and beta chains (epsilon only when 0-6 weeks of prenatal age)
Time frames for gamma, beta and delta chain transcription in the beta chains for hemoglobin
Gamma: 2 weeks prenatal -> 30 years
peaks at 18 weeks prenatal
Beta: 7 weeks prenatal -> life
* peaks at 18 years*
Delta: 30 weeks -> 40 years
* very low levels throughout though*
Why are gamma chains present at such Huge levels when prenatal vs postnatal?
HbF (2 alpha and 2 gamma chains) has a higher affinity to oxygen and doesn’t bind as well to 2,3 BPG
- 2,3 BPG is used to help facilitate transfer of oxygen from maternal circulation to the fetal circulation
HbF is preferred over HbA (normal) at this stage
Groups of hemoglobinopathies
Qualitative: structural defects
Quantitative: defects in total synthesis
What is the most common structural defects in hemoglobin?
Point mutations in the B-globin gene
Sickle cell= glu -> Val
HbC = glu -> lys
Treatment of sickle cell disorder
Hydration, analgesics, antibody therapy
- hydroxyurea (increases circulating levels of HbF and reduces RBC sickling)
- stem cell transplantation is possible in severe cases
Treatment of Methemoglobinemia
Methylene blue dye (acts as electron acceptor and helps reduce iron beck to Fe2+/normal state)
Vitamin c is also used in conjunction with methylene blue to speed up this process
Selective advantage against malaria
Homozygous individuals for malaria are much more likely to die than heterozygote individuals
in fact, heterozygous individuals have increases resistance to some strains (such as plasmodium falciparum malaria).
- because of this, the trait of sickle cell anemia is actually selected for in cultures with high rates of malaria (such as Africa)
Results in disease maintaining a high frequency in populations that are often exposed to malaria, but not in populations with low exposure to malaria
Glutathione (GSH) structure
Glycine, cysteine and glutamate residues bound together and have a sulfide binding site
Types of G6PD deficiencies
Class 1: very severe (<10% activity)
Class 2: severe (<10% enzyme
activity)
- CGPD Mediterranean
Class 3: moderate (10-60% enzyme activity)
- G6PD A-
Class 4: asymptomatic (60% enzyme activity)
Mutations can affect the stability active site or allosteric site of G6PD.
- most common is overall stability
Nutritional anemia types
Microcytic anemia (MCV <80) - often signals deficency in iron, Vit. C or pyridoxine
Normocytic (MCV = 80-100
- protein malnutrition
Macrocytic (MCV >100)
- often signals deficency in vitamin B12 and/or folic acid
