KINETICS AND EQUILIBRIUM OF THE REACTION OF ELLMAN'S REAGENT WITH DOMESTIC HORSE (Equus caballus) HAEMOGLOBIN

📄 Project Abstract

CysF9[93] exists in two tertiary conformations, r and t, which are in dynamic equilibrium. The reactivity of the CysF9[93] sulphydryl group and the oxygen affinity of haemoglobin (Hb) are affected by protons (H+) and organic phosphates such as inositol hexakisphosphate (inositol-P6). This study was aimed at determining the effects of inositol-P6 and pH on the relative populations of the two conformations. The major and minor haemoglobins in horse haemolysate were separated using a column of Whatman's carboxymethylcellulose (CMC 52). Equilibrium studies of the reaction of CysF9[93] with Ellman's reagent, 5,5'-dithiobis(2-nitrobenzoate) (DTNB), equilibrated for six hours in the presence and absence of inositol-P6 were carried out at 25 degrees C. The absorbance of the product of DTNB reaction, 5-thio-2- nitrobenzoate, at different concentrations of DTNB and 50 mol (haem) dm-3 was read at 412 nm. The absorbance was then substituted into an appropriate equation derived for the determinatio...