Determination of inorganic phosphate by coupling thymidine phosphorylase and reversed-phase high-performance liquid chromatography: Application to tonoplast pyrophosphatase activity

被引：12

作者：

Hill, M ^{[1
]}

Arrio, B ^{[1
]}

机构：

[1] Univ Paris Sud, ERS 571 CNRS, Ctr Orsay, F-91405 Orsay, France

来源：

ANALYTICAL BIOCHEMISTRY | 1997年 / 254卷 / 02期

关键词：

D O I：

10.1006/abio.1997.2437

中图分类号：

Q5 [生物化学];

学科分类号：

071010 ; 081704 ;

摘要：

We developed an HPLC method for the measurement of inorganic phosphate using thymidine phosphorylase (EC 2.4.2.4). This enzyme catalyzes the phosphorolysis of thymidine to a-deoxyribose l-phosphate and thymine. Thymine release was measured at 265 nm after separation by reverse-phase HPLC. The assay was sensitive enough to detect as little as 10 pmol of Pi. The response to the phosphate concentration was linear from 1 to 100 mu M. The value of this method was demonstrated in an analysis of the kinetics of Pi release from PPi in the presence of Catharanthus roseus tonoplast pyrophosphatase. (C) 1997 Academic Press.

引用

页码：249 / 253

页数：5

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