Purification and properties of 2,3-bisphosphoglycerate phosphatase-mutase from erythrocytes of day-old chicks

1 Large quantities of 2,3‐bisphosphoglycerate accumulate in the red blood cells of the chick embryo during the week prior to hatching; the 2,3‐bisphosphoglycerate then abruptly decreases to very small amounts within a few days after hatching. 2 The enzyme 2,3‐bisphosphoglycerate phosphatase was purified from the red blood cells of the day old chick. The elution profiles for bisphosphoglycerate phosphatase and 2,3‐bisphosphoglycerate mutase were identical upon gradient elution from columns of hydroxylapatite and diethylaminoethylcellulose and by gel filtration on Sephadex G‐100. At each stage of purification the ratio of phosphatase to mutase activity was the same. It was concluded that, just as in the human erythrocyte, both bisphosphoglycerate phosphatase and 2,3‐bisphosphoglycerate mutase activities in chick erythrocytes reside on one protein. 3 Bisphosphoglycerate phosphatase has a pH of optimal activity of 7.0. Its activity is stimulated by 2‐phosphoglycolate, phosphoenolpyruvate, bisulfite and dithionite and is inhibited by 2‐phosphoglycerate, 3‐phosphoglycerate, inorganic pyrophosphate and phytic acid. These properties are very similar to those described for bisphosphoglycerate phosphatase purified from human erythrocytes. 4 The purified bisphosphoglycerate phosphatase‐mutase contained 3‐phosphoglycerate mutase activity. Although the 2,3‐bisphosphoglycerate mutase and 3‐phosphoglycerate mutase activities were nearly equal in the purified enzyme, this protein accounts for a maximum of only 6% of the 3‐phosphoglycerate mutase activity in these cells. 5 The total activity of the bisphosphoglycerate phosphatase in the erythrocytes was measured at intervals during development of the embryo, in the young chick, and in the mature chicken. Its activity increases during the time of 2,3‐bisphosphoglycerate accumulation. The enzyme activity decreases gradually from its maximum in the two‐day‐old chick (1.67 μmol h−1 g hemoglobin−1) and by 27 days it has fallen nearly to the levels seen in the adult chicken (0.11 μmol h−1 g hemoglobin−1). 6 The factors which might be involved in inducing the 2,3‐bisphosphoglycerate accumulation and degradation in the late chick embryo and young chick are discussed. It is concluded that this system is very similar to that found in the erythrocytes of man.