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The preparation and assay of sedoheptulose-7-phosphate

1970; Elsevier BV; Volume: 141; Issue: 2 Linguagem: Inglês

10.1016/0003-9861(70)90160-8

1096-0384

T. Wood, Wilson Poon,

Erythrocyte Function and Pathophysiology

The enzymic synthesis of sedoheptulose-7-phosphate from ribose-5-phosphate in the presence of ribose-5-phosphate ketol-isomerase, ribulose-5-phosphate-3-epimerase, and transketolase was investigated. The reaction was driven in the direction of heptulose formation by the addition of NAD, glyceraldehyde phosphate dehydrogenase, and arsenate. These three enzymes could be replaced by a liver supernatant. The sedoheptulose-7-phosphate was purified by barium precipitation and separated from arsenate by gradient elution from a column of DEAE-Sephadex A-25. The product was assayed colorimetrically with cysteine-sulfuric acid and enzymically with transaldolase and by a new procedure. The new enzymic procedure involved the, phosphorylation of the monophosphate to sedoheptulose-1,7-diphosphate by ATP and phosphofructokinase and the cleavage of the diphosphate by aldolase to erythrose-4-phosphate and dihydroxyacetone phosphate. In the presence of glycerol-1-phosphate dehydrogenase the latter was reduced to glycerol-1-phosphate with the oxidation of an equivalent amount of NADH.