Equilibrium Studies of the Reaction of Turkey (Meleagris gallopavo) Haemoglobin Sulphydryl Groups with 5,5′-dithiobis(2-nitrobenzoate): Tertiary Conformational Change in Turkey Haemoglobin Induced by Inositol hexakisphosphate

Abstract

The red blood cell of turkey contains two haemoglobin types, major and minor components. In the present study, the equilibrium constant, Kequ, for the reaction of 5,5′-dithiobis(2-nitrobenzoate), DTNB, with the sulphydryl group of the major turkey aquomethaemoglobin was determined at 25°C as a function of pH. Kequ varies by about 2 to 3 orders of magnitude between pH 5.6 and 9.0 for both haemoglobin [stripped and in the presence of inositol hexakisphosphate (inositol-P6)]. Calculations from the pH dependence of Kequ showed that in the r ⇌ t tertiary conformational transition of aquomethae-moglobin, the t isomer population was 0.26 %. In the presence of inositol-P6, the t isomer population increased to 9.08 %. The results showed that while inositol-P6 increased the relative population of the t tertiary conformation by changing the relative distribution of two protein conformations, it had no effect on Kequ. The effect of Inositol-P6 on the nature and number of groups linked to the DTNB reaction was also determined.