OXYGENATION PROPERTIES OF HUMAN ERYTHROCYTES CONTAINING EXCLUSIVELY α-NITROSYL HEMOGLOBIN: A PROMISING BLOOD TRANSFUSANT CANDIDATE

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Abstract
We have prepared human erythrocytes that contain exclusively alpha-nitrosyl hemoglobin (Hb), i.e., alpha(Fe-NO)2beta(Fe-O2)2, by incorporating nitric oxide (NO) into erythrocytes in a well-controlled nitrosylation process. The amount of alpha(Fe-NO) corresponding to 50% of the total heme content of the erythrocytes and exclusive binding of NO to alpha-subunits of intraerythrocytic Hb were confirmed by EPR. Oxygenation experiments on the intraerythrocytic alpha-nitrosyl Hb over a wide range of pH showed that: (1) the oxygen affinity of cell-free and intraerythrocytic alpha-nitrosyl Hbs were much lower than native Hb in their respective environments; (2) the oxygenation characteristics of the intraerythrocytic alpha-nitrosyl Hb in the acidic range was similar to that of the cell-free alpha-nitrosyl Hb in the presence of 2,3-diphosphoglycerate; and (3) the apparent Bohr effect in the intraerythrocytic alpha-nitrosyl Hb was dramatically diminished. This can be due to a restricted variation in intraerythrocytic pH in the alkaline region and the presence and/or production of endogenous 2,3-diphosphoglycerate. By comparing oxygen saturation characteristics, it was found that the intraerythrocytic alpha-nitrosyl Hb, despite its halved oxygen carrying capacity, could deliver more oxygen than DPG-depleted erythrocytes under similar experimental conditions. This makes alpha-nitrosyl Hb-containing erythrocytes a promising candidate for blood transfusant.