Enzymatic assay of inorganic phosphate with use of sucrose phosphorylase and phosphoglucomutase

M. Tedokon, K. Suzuki, Y. Kayamori, S. Fujita, Y. Katayama

研究成果: Contribution to journalArticle

26 被引用数 (Scopus)

抄録

We developed a new enzymatic method for the assay of inorganic phosphate (Pi) by using sucrose phosphorylase (SP; EC 2.4.1.7) and phosphoglucomutase (PGM; EC 5.4.2.2). Pi is transferred to sucrose by SP, producing α-D-glucose 1-phosphate (G1P) and α-D-fructose. G1P is transphosphorylated by PGM in the presence of α-D-glucose 1,6-bisphosphate to form α-D-glucose 6-phosphate, which is oxidized by NAD+ and glucose-6-phosphate dehydrogenase (EC 1.1.1.49) to form 6-phosphogluconate (6PG) and NADH. Finally, the oxidation of 6PG by NAD+, catalyzed by 6-phosphogluconic dehydrogenase (EC 1.1.1.44), yields D-ribulose 5-phosphate and NADH. Thus two molecules of NADH are formed for each molecule of Pi, and the reaction is monitored at 340 nm. The K(m) values of SP for Pi and sucrose were 4.44 and 5.31 mmol/L, respectively. The best buffer was 1,4-piperazinediethanesulfonic acid (PIPES) at 50 mmol/L and pH 6-7. Implementing this method with a Cobas-Bio centrifugal analyzer allowed us to measure Pi accurately and precisely.

本文言語英語
ページ(範囲)512-515
ページ数4
ジャーナルClinical Chemistry
38
4
DOI
出版ステータス出版済み - 1992

All Science Journal Classification (ASJC) codes

  • Clinical Biochemistry
  • Biochemistry, medical

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