Identification of residues contributing to the Bohr effect of human haemoglobin

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We have searched for the amino acid residues responsible for the acid Bohr effect and for that part of the alkaline one which had not been accounted for by Vall α, HisH5(122)α and HisHC3(146)β. We have measured the Bohr effect by titrating the protons released or taken up on oxygenation by the haemoglobins of various species and by various abnormal haemoglobins. We have identified HisH21(143)β as the residue responsible for about half the acid Bohr effect observed in stripped haemoglobin in 0·1 m-KCl at 25°C. Under the same conditions all abnormal haemoglobins in which LysEF6(82)β is replaced by a neutral or anionic residue have reduced alkaline Bohr effects. This reduction is strongly dependent on chloride concentration. The combined evidence obtained by ourselves and others indicates that in stripped haemoglobin in 0·1 m-chloride LysEF6(82)β is responsible for the missing part of the alkaline Bohr effect, and that this is also the part that is due to weakly bound chloride, while Vallα accounts for the part of the alkaline Bohr effect that is due to strongly bound chloride. Since LysEF6(82)β is also one of the residues binding d-glycerate 2,3-bis(phosphate), that part of the alkaline Bohr effect in stripped haemoglobin for which it is responsible would be of no significance in vivo.

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