一种用于检测δ1-吡咯啉-5-羧酸合成酶活性的放射性同位素测定法。

A radioisotopic assay for delta 1-pyrroline-5-carboxylate synthase activity.

作者信息

Smith R J

出版信息

Enzyme. 1984;31(2):115-21. doi: 10.1159/000469510.

Abstract

A radioisotopic assay is described for measuring the activity of delta 1-pyrroline-5-carboxylate synthase, the enzyme that catalyzes the formation of delta 1-pyrroline-5-carboxylic acid from glutamic acid. Pyrroline-5-carboxylic acid is a common intermediate in the pathways through which glutamic acid, proline, and ornithine are interconverted. To determine pyrroline-5-carboxylate synthase activity, cell homogenates are incubated with [14C]-glutamic acid, the products of the reaction are converted quantitatively to proline by sodium borohydride, and proline is isolated by cation-exchange column chromatography. Cofactor requirements have been defined, and the activity of pyrroline-5-carboxylate synthase in several different cultured fibroblast lines is reported.

摘要

本文描述了一种放射性同位素测定法，用于测量δ1-吡咯啉-5-羧酸合成酶的活性，该酶催化由谷氨酸形成δ1-吡咯啉-5-羧酸。吡咯啉-5-羧酸是谷氨酸、脯氨酸和鸟氨酸相互转化途径中的常见中间体。为了测定吡咯啉-5-羧酸合成酶的活性，将细胞匀浆与[14C]-谷氨酸一起孵育，反应产物用硼氢化钠定量转化为脯氨酸，然后通过阳离子交换柱色谱法分离脯氨酸。已经确定了辅助因子的需求，并报道了几种不同培养的成纤维细胞系中吡咯啉-5-羧酸合成酶的活性。