Contrasting behavior of oxy- and carbonmonoxyhemoglobin: reaction of Ellman’s reagent with CysF9[93]β of carbonmonoxyhemoglobin reveals tertiary level conformational differences between mammalian hemoglobins

Abstract

The efficiency of oxygen transport function of hemoglobin is affected by their amino acid sequences amongst other things. To gauge how the amino acid sequences of mammals affect the affinity of their hemoglobins for oxygen, the pH dependent studies of the affinities of CysF9[93]β sulfhydryl group of oxy- and carbonmonoxyhemoglobin of four mammalian hemoglobins: rabbit, straw coloured fruit bat (SCFB), human A (HHbAA) and human S (HHbSS) for 5,5’-dithiobis(2-nitrobezoate) (DTNB) in dilute solution were reported. The affinities of DTNB for the hemoglobins in each of the derivatives decreases with increasing pH. Rabbit hemoglobin has an affinity that is at least twice that of HHbSS for DTNB. That of SCFB and HHbAA which were quite comparable were at least three times the affinity HHbSS at corresponding pH. The proportion of r-conformation in the oxy derivative of all the mammalian hemoglobins were between 73 and 80 % at high pH. The proportion of r-conformation in the carbonmonoxy derivative were 0.62% for SCFB; 8.0% for rabbit; 88.60 % for HHbAA and 91.87% for HHbSS. Carbonmonoxyhemoglobin is more sensitive to increasing differences in amino acid sequence and composition of hemoglobin compared to that in oxy-derivative. The restriction in the isomerization process of oxyhemoglobin was rationalized on the basis of ease of formation of hydrogen bond between O and N of α and β subunit by HisE7[63]β in oxyhemoglobin compared to COHb where such linkages are hindered. Single point mutation in human hemoglobin at position 6β result in significant difference in affinity of CysF9[93]β for DTNB.