Multifunctionality of the Enzyme Involved in the 2,3-Diphosphoglycerate Metabolism of Pig Erythrocytes

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Abstract
Diphosphoglyceromutase, 2,3-diphosphoglycerate phosphatase and phosphoglyceromutase were simultaneously purified from pig erythrocytes. Three fractions (peaks I, II and III) which had all 3 activities in different ratios were obtained by column chromatography. Peak I was extremely rich in phosphoglyceromutase, containing more than 90% of the total activity of this enzyme. In contrast, peak III was active in metabolizing 2,3-diphosphoglycerate, containing about 95% of both the diphosphoglyceromutase and the 2,3-diphosphoglycerate phosphatase activity. It seems likely that peak I functions as phosphoglyceromutase and that peak III functions in the 2,3-diphosphoglycerate metabolism. The homogeneity of peak III was established by disc gel electrophoresis in the presence and the absence of sodium dodecyl sulfate, as well as by ultracentrifugation. The 3 activities of peak III were lost at the same rate on thermal inactivation. The 2 enzyme activities for metabolizing 2,3-diphosphoglycerate, which were believed to be due to different proteins, are probably attributable to 1 protein, along with some phosphoglyceromutase activity. The diphosphoglyceromutase activity of this protein was inhibited by the product, 2,3-diphosphoglycerate, as well as by hydroxypyruvate phosphate, 2-phosphoglycolate, inorganic phosphate and bisulfite. The 2,3-diphosphoglycerate phosphatase activity was enhanced by 2-phosphoglycolate and hydoxypyruvate phosphate, and by the coexistence of chlorine ion and bisulfite, while it was inhibited by monophosphoglycerates. The native peak III protein had a MW of 59,000 as determined by equilibrium centrifugation. Disc gel electrophoresis in the presence of sodium dodecylsulfate yielded a single protein band with a MW of 28,000, indicating that this protein was composed of 2 subunits with a similar molecular weight. Occurrence of multifunctional proteins in pig erythrocytes is compared with that in human erythrocytes from the standpoint of the universal existence of such proteins.

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