Purification and Properties of 2,3-Bisphosphoglycerate Phosphatase-Mutase from Erythrocytes of Day-Old Chicks

Abstract

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.