Okonjo, K. O. and Fodeke, A. A.
Reversible reaction of 5,5V-dithiobis(2-nitrobenzoate) with the hemoglobins of the domestic cat: Acetylation of NH3
+ terminal group of the h chain transforms the complex pH dependence of the forward apparent second order rate constant to a simple form.
We demonstrate kinetically that the reaction of 5,5V-dithiobis(2-nitrobenzoate) with the CysF9h sulfhydryl group of domestic cat hemoglobins is a reversible process. In the major hemoglobin, in which the NH3 + terminal group of GlyNA1h is free, kf, the apparent
forward second order rate constant, has a complex pH dependence profile. In the minor hemoglobin, the NH3
+ terminal group of SerNA1h is acetylated, and the pH dependence profile of kf is simple. These results support the proposal that the positively charged groups at the organic phosphate binding site are electrostatically linked to CysF9h. Quantitative analyses of the complex profiles enabled us to estimate pKas of 7.47T0.3; 6.53T0.03 and 8.49T0.3 for GlyNA1h, HisH21h and other histidines within 2 nm of the sulfhydryl, and CysF9h,
respectively, of the major hemoglobin. Analyses of the simple profiles gave pKas of 6.33T0.17 and 8.54T0.5 for HisH21h and other histidines within a distance of 2 nm of the sulfhydryl, and CysF9h of the minor hemoglobin, respectively.
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