Electrochemical modulation of sickle cell haemoglobin polymerisation.
Sickle cell haemoglobin (HbS) differs from normal haemoglobin by a single amino acid in its beta chain. This amino acid replacement, from glutamic acid to valine, causes polymerisation of proteins into defined long insoluble fibres with a typical diameter of 21.5 nm. The polymerisation is triggered by the formation of deoxyhaemoglobin (deoxyHb) from oxyhaemoglobin (oxyHb) in low oxygen partial pressures, which results in a conformational change in the secondary structure of the protein. We describe an electrochemical method to modulate the oxygen concentration in an optically transparent thin layer cell to produce deoxyhaemoglobin whilst monitoring the extent of polymerisation using turbidity measurements. The oxygen is depleted in the vicinity of the electrode and triggers the polymerisation. The kinetics of polymerisation were investigated using a model for fibrillogenesis describing a two-step process of nucleation followed by elongation. Rate constants describing the nucleation and growth at monomer concentration of 300 mg cm(-3) (4.65 x 10(-3) M) were determined to be 9.45 (+/-0.08) x 10(-6) s(-1) and 1.22 (+/-0.03) x 10(-3) s(-1) respectively, showing that nucleation was far slower than the growth. A similar difference between the rate constants for the nucleation (2.99 (+/-0.4) x 10(-8) s(-1)) and growth (1.08 (+/-0.2) x 10(-3) s(-1)) was seen at monomer concentration of 50 mg cm(-3) (7.75 x 10(-4) M). These results show that nucleation was monomer concentration dependent; however growth was largely independent of monomer concentration. In this study we present a methodology that may be used as a screening method for substances that effect the fibre nucleation and or growth that could be valuable to the pharmaceutical industry for treating sickle cell disease.
Iqbal
Zeshan
Z
Department of Chemistry, University College London, 20 Gordon St., London, WC1 H 0AJ, UK.
McKendry
Rachel
R
Horton
Michael
M
Caruana
Daren J
DJ
D000755:Anemia, Sickle Cell / Q000097:blood*; D001704:Biopolymers; D004563:Electrochemistry / Q000295:instrumentation / Q000379:methods; D006451:Hemoglobin, Sickle / Q000737:chemistry*; D006454:Hemoglobins / Q000737:chemistry*; D006801:Humans; D010108:Oxyhemoglobins / Q000737:chemistry*
D001704:Biopolymers; D006451:Hemoglobin, Sickle; D006454:Hemoglobins; D010108:Oxyhemoglobins; C100003:deoxyhemoglobin