Partial characterization of the inactive mutant form of human red cell bisphosphoglyceromutase and comparison with an alkylated form

• Published in: Biochimica et biophysica acta Vol. 742; no. 1; pp. 243 - 249
• Main Authors: Rosa, Raymonde, Préhu, Marie-Odette, Albrecht-Ellmer, Krystina, Calvin, Marie Claude
• Format: Journal Article
• Language: English
• Published: Netherlands Elsevier B.V 12.01.1983
• Subjects: Alkylation; Animals; Antibody; Antigen-Antibody Complex; Bisphosphoglycerate Mutase - blood; Bisphosphoglycerate Mutase - genetics; Bisphosphoglycerate Mutase - isolation & purification; Bisphosphoglyceromutase; Chickens - immunology; Drug Stability; Erythrocytes - enzymology; Hot Temperature; Human red cell; Humans; Immunodiffusion; Immunoelectrophoresis; Inactivation; Kinetics; Mutant enzyme; Mutation; Phosphotransferases - genetics; Substrate binding; Mutant enzyme; Antibody; 2,3-DPG; Inactivation; Bisphosphoglyceromutase; Substrate binding; Human red cell
• ISSN: 0167-4838; 0006-3002; 1879-2588
• DOI: 10.1016/0167-4838(83)90382-5

The trifunctional enzyme bisphosphoglyceromutase (or diphosphoglycerate mutase) (EC 2.7.5.4) was purified from human red cells and injected into two chickens. Specific anti-bisphosphoglyceromutase antibodies were produced that displayed a single precipitation line on Ouchterlony plates and on immunoelectrophoresis. No cross-reaction of these antibodies was detected with phosphoglyceromutase, the common glycolytic enzyme. Immunoneutralization of bisphosphoglyceromutase and of its two other activities, i.e., bisphosphoglycerate phosphatase and phosphoglyceromutase, was observed for a purified preparation. The anti-bisphosphoglyceromutase antibody reacts with the inactive enzyme present in the hemolysate of a mutant human subject. It also binds bisphosphoglyceromutase inactivated by N-ethylmaleimide, a strong alkylating agent of SH groups. Active bisphosphoglyceromutase is stable at 55°C, whereas the inactive forms of the mutant and of the alkylated hemolysates are thermolabile. These forms can be protected against thermal precipitation by 4 mM 2,3-diphosphoglycerate and 4 mM 3-phosphoglycerate. These findings afford evidence that the binding of the substrates on the bisphosphoglyceromutase molecule is not prevented by alkylation nor by the mutation of the hereditary inactive enzyme.