Hemoglobin Variants

Question 1

most Hb has what subunits?

Answer 1

a₂B₂

Question 2

Hb can be composed of what polypeptides?

Answer 2

alpha, beta, gamma, delta

Question 3

what chromosomes and gene clusters produce the polypeptides (chain) assembled into various hemoblogins?

Answer 3

• chromosomes 11 and 16
  
  • chromosome 11
    
    • alpha like cluster –>
      
      • alpha chains
      • zeta chains
  • chromosome 16
    
    • beta like gene cluster –>
      
      • beta chains
      • gamma chains
      • delta chains
      • epsilon chains

Question 4

discuss the Hb variants present throughout development, and what

• poypeptides they consist of
• genes those polypeptides came from
• chromsomes those genes are on

Answer 4

• embryonic = alpha, gamma, zeta and episilon polypeptides used.
  
  • zeta (from alpha cluster) and episilon (from beta cluster) are ONLY seen in embryonic.
  • combos:
    
    • zE (gower 1)
    • aE (gower 2)
    • portland (zy)
• fetal = a₂y₂
• adult = a₂B₂ > a₂d₂

Question 5

the globin genes are activated in what direction along the chromosome? what effect does this have?

Answer 5

5’ –> 3’ direction.

• polypeptides close to the 5’ end seen in earlier in life (zeta, episilon, gamma) and the 3’ end seen later in adulthood (beta, delta)

Question 6

what leads to hereditary persistence of fetal Hb? what are the clinical imlications of this condition?

Answer 6

• usually due to improper expression/deletion of the beta and delta globin genes, which are:
  
  • found closer to the 3’ of their chromosomes & normally expressed only in adulthood.
    
    • as a result, embryonic/ fetal chains (z, E, y) persist in the blood.
      
      • sustained levels of fetal Hb can be useful:
        
        • sickle cell
        • B-thalassemia

Question 7

discuss the solubility of Hb & its significance

Answer 7

• the Hb tetramer is highly soluble
• however, individual subunits are not.
  
  • so, is subunits are not expressed in sufficient amounts/improperly assembled they can precipiate –> inclusions

Question 8

discuss the primary, secondary, tertiary & quaternary strucutres of globins

Answer 8

• primary: aa sequence - highly similar for each globin (polypeptide)
• secondary: rich in alpha helices
• tertiarty: hyrophobic aas in core, hydrophillic aas out
• quaternary: two aB dimers (or other variation)

Question 9

Hb quaternary structure is highly dependent on what interactions?

Answer 9

1. proper hydrophobic/hydrophillic aa alignment
2. a₁b₁ & a₁B₂ contant points
3. His residue in F helix

Question 10

how many amino acids are there?

Answer 10

20

Question 11

what type of mutation causes sickle cell anemia (HbS)?

Answer 11

point mutation:

• Glu –> Val in B subunit
  
  • this is a loss ot two negative charges:
    
    • Glu = -, hydrophillic
    • Val = neutral, hydrophobic

Question 12

discuss the pathogenesis of sickle cell anemia

Answer 12

• Glu –> Val mutation.
• This Val residue is on the surface of the T-state (tense) molecule
• Val residue interacts with other hydrophobic aas (phenylalinine, leucine) –> Hbs molecules aggregate
  
  • 14 HbS total HbS fiber

Question 13

why is HbS aggregates less likely to form when HbS is oxygenated?

Answer 13

• when oxygenated form, HbS is the in R-state.
  
  • the arrangement of aas shifts such that key hydrophobic residues (F, L) available to interact with mutant Val in the T-state become buried in the HbS core.
  • Val cannot make the non-polar bonds that lead to aggregation & HbS fiber formation.

Question 14

how to dx sickle cell anemia?

Answer 14

• by electrophoresis
  
  • due to structural HbS migrates differently than normal adult forms of HbA - a₂B₂, a₂D₂
    
    • the abnormal beta subunit (S)

Question 15

heinz bodies

• defintition
• what forms them?
• how are they detected?
• clinical significance?

Answer 15

• inclusions of denatured, aggregated Hb (typically Hb variants) within RBCs
• clinical:
  
  • congenital heinz body hemolytic anemia (CHBA) - these RBCS are more susceptible to hemolysis
  • dx - smears stained for reticulocytes

Question 16

discuss the clinical effects of high vs low affinity Hb variants

Answer 16

• high affinity: decreased O₂ delivery
  
  • asympomatic (m/c)
  • familial erthrocytosis
• low affinity: normal O2 deliver, low O2 saturation
  
  • asymptomatic +/- cyanosis

Question 17

M hemoglobin

• definition
• clinical significance

Answer 17

• Hb variation seen in methemoglobinema (ferric iron)
• clinical presentation
  
  • asymptomatic, since patients has metHb reductase
    
    • lavendar blue skin, chocolate brown blood

Question 18

alpha thalassemia

• cause/pathogenesis
• clinical significance

Answer 18

• cause/pathogenesis:
  
  • disruption of the all four alleles for alpha chain (chromosome 16)
  • a chain not made in suffcient quantity
    
    • HbH = 4 B-subunits
      
      • NO COOPERATIVITY
      • ​HIGH AFFINITY
• clinical: poor oxygen release

Question 19

beta thalassemia

• cause, pathogenesis
• clinical significance

Answer 19

• cause/pathogenesis
  
  • disruption of two alleles for Beta chain (chromsome 11)
  • only alpha chains produced
    
    • a subunits are insoluble –> precipitate in RBCs –> hemolysis –> Cooley anemia

Question 20

what is AHSP?

why is it important?

how does it work?

Answer 20

• alpha-hemoblogin stabilizing protein
  
  • forms a soluble complex with newly synthesized, free alpha chain monomers preventing their aggregation/misfolding
    
    • ​binds both oxy & deoxy a-chain
      
      • the a-AHSP interface & aB interface in Hb is indentical
      • as B subunits are made, they displace ASHP
  • this is key since there are 4 a-chain alleles vs 2 B chain-alleles, & we need a way to manage excess a-subunits

Question 21

how does the structure of the alpha-chain change when it is bound to ASHP

Answer 21

this only applies to an oxygenated subunit:

• the Fe²⁺ atom in that subunit ecomes bound to the distal histine rather than the proximal histidine

Question 22

ATXR

• definition
• inheritance pattern
• clinical significance

Answer 22

• alpha thalaessema X-inked intectual disability syndrome
• X-linked recessive
• characterized by intellectual disability & affects multiple organs

Question 23

what is HbH?

Answer 23

• a hemoblogin variant made of only B-subunits (a homotetramer)
  
  • seen in alpha-thalasemmia
  • HbH = high oxygen affinity + no cooperativity = poor oxygen delivery

Question 24

HbCS mutation

• cause/pathogenesis
• clinical significance
• treatment

Answer 24

• mutation at the termination codon of the a2-globin gene
  
  • causes abnormally long alpha chains
• clinical –> can lead to thallasemia intermedia
  
  • ​delayed growth
  • weak bones
  • enlarged sleen
• tx - blood transufion

Question 25

thalasemia intermedia?

Answer 25

* due to HBCS hemoglobin variant * mutation in a2 subunit, specically, which makes this alpha chain too long * --\> growth defects: abrnomal bones, enlarged spleen