EFFECT OF A3[6]BETA(GLU-]VAL) MUTATION ON REACTIVITY OF THE CYSF9[93]BETA SULFHYDRYL-GROUP OF HUMAN HEMOGLOBIN-S

The pH dependence profiles of the apparent second-order rate constant, k(app), for the reaction of the oxy, carbon monoxy and aquomet deriva tives of human haemoglobin S with 5,5'-dithiobis(2-nitrobenzoate) (DTN B) in buffers of ionic strength 50 mmol dm(-3) are complex. The pK(a) s of the ionizable organic phosphate binding groups which influence th e reactivity of the CysF9[93]beta sulfhydryl group have been determine d from quantitative analyses of the complex profiles. These pK(a) valu es were not significantly different from those of haemoglobin A. In th e presence of inositol hexakisphosphate (inositol-P-6) each profile as sumes a simple form resembling the titration curve of a diprotic acid. The pK(a) s of HisHC3[146]beta and CysF9[93]beta were determined from quantitative analyses of the simple profiles. The mean values obtaine d, 6.6 +/- 0.2 and 8.84 +/- 0.04, respectively, were the same as those of haemoglobin A. Comparison of the k(app) data for haemoglobin S wit h those of haemoglobin A shows that, irrespective of the presence or a bsence of inositol-P-6 presence, the carbon monoxy and aquomet derivat ives of haemoglobin A react more rapidly than the corresponding haemog lobin S derivatives. In contrast, in the absence of inositol-P-6, oxyh aemoglobin A reacts faster than oxyhaemoglobin S; in the presence of t he organic phosphate both haemoglobins react at about the same rate. A t an ionic strength of 200 mmol dm(-3) in the absence of inositol-P-6, the pH dependence profiles of k(app) for the oxy, carbon monoxy and a quomet derivatives of haemoglobins A and S are simple. Quantitative an alyses of these profiles give mean pK(a) values of 5.4 +/- 0.1 and 8.9 +/- 0.2 for HisHC3[146]beta and CysF9[93]beta, respectively. The oxy and carbon monoxy derivatives of both haemoglobins react at about the same rate, but aquomethaemoglobin A reacts significantly more rapidly than aquomethaemoglobin S.