2010B15 Explain the role of haemoglobin as a buffer.

· Intro: Hb, buffer

· Hb buffering mechanism

· Hb buffering efficiency

Haemoglobin

· Specialised O₂ carrying protein in RBCs

· 4 x Haem: protoporphyrin ring and Fe²⁺

· 4 x Globin polypeptide chain (2α, 2β)

· Important buffer in ECF

Buffer

· Substance resisting ↓/↑ pH when acid/base added to solution

· = weak acid and conjugate base

· i.e. H⁺ + B^- <-> HB

Effective buffer

· Abundant

· Rapid

· pKa near prevailing pH (+/- 1)

· Open-ended (i.e. carbonic acid system – CO₂ excreted by lungs, HCO₃^- and H⁺ by kidneys)

Diffusion	· Gases pass down partial pressure gradients · O₂: RBC -> tissue · CO₂: tissue -> RBC
H⁺ buffering	· · H⁺ + KHb <-> HHb + K⁺ (pKa 6.8) · Active component is imidazole groups of histidine residues · HCO₃^- exchanged for Cl^- across the RBC membrane (=Hamburger effect)
Carbamino formation	· -NH₂ + CO² <-> -NHCOO^- + H⁺ · i.e. binds to terminal amino groups and amino groups on side chains of arginine and lysine · H⁺ buffered as above
Haldane effect	· DeoxyHb takes up more H⁺ and CO₂ · Minimises ↓pH from arterial (7.44) to venous (7.36) despite large ↑n(H⁺) · 70% of effect: due to 3.5x ↑formation of carbaminoHb · 30% of effect: imidazole pKa 6.6 -> 8.2 hence ↑buffering · For every mmol O₂ unloaded, 0.7mmol H⁺ is loaded without ↓pH (= isohydric buffering)

(note: reverse changes occur at the lungs, facilitating O₂ uptake and CO₂ excretion)

High quantity	· High [Hb] (140g/L) (cf. plasma proteins 70g/L) · Large number imidazole groups per Hb (39) (cf. plasma proteins 13)
pKa near pH	· pKa 6.8 (oxyHb 6.6, deoxyHb 8.2) · pH 7.34-7.45
Open-ended	· Indirectly via link to carbonic acid system
Rapid	· CO₂ lipid soluble, rapidly enters RBC · Carbonic anhydrase very fast (10,000x / second)