Transition of haemoglobin between two tertiary conformations: Inositol hexakisphosphate increases the transition...

The equilibrium constant (K(equ)) for the reaction of 5,5'-dithiobis(2-nitrobenzoate) - DTNB - with the CysF9[93]beta sulphydryl group of the haemoglobins of the sheep decreases by about two orders of magnitude between pH approximately 5.6 and 9.2: from a mean of 7.2+/-1 to a mean of 0.044+/-0.01. Calculations from the pH dependence of K(equ) show that in the r-t tertiary conformational transition of haemoglobin the t isomer population is 50.7 and 61.8% for the major and minor haemoglobins, respectively. In the presence of inositol hexakisphosphate (inositol-P(6)), K(equ) increases for both haemoglobins by about an order of magnitude through most of the pH range. The t isomer population also increases to 82.1 and 79.6% for the major and minor haemoglobins, respectively. These results indicate that inositol-P(6) increases the affinity of the sulphydryl for DTNB by increasing the population of the t isomer. It is highly probable that a minimum four-state model that includes the r-t transition is required for a full understanding of haemoglobin function.