采用高效液相色谱荧光检测法测定牛松果体中血清素-N-乙酰转移酶的动力学特性。

Determination of kinetic properties of serotonin-N-acetyltransferase in bovine pineal gland using HPLC with fluorimetric detection.

作者信息

Fajardo N, Abreu P, Alonso R

机构信息

Departamento de Fisiología, Facultad de Medicina, Universidad de La Laguna, Tenerife, Spain.

出版信息

J Pineal Res. 1992 Sep;13(2):80-4. doi: 10.1111/j.1600-079x.1992.tb00058.x.

DOI:10.1111/j.1600-079x.1992.tb00058.x

PMID:1453312

Abstract

The determination of serotonin-N-acetyltransferase (NAT) activity in the bovine pineal gland and other rat tissues was based upon the separation and detection of N-acetyltryptamine formed from tryptamine and acetyl CoA by means of high performance liquid chromatography with fluorimetric detection. In the bovine pineal the enzyme exhibited a Km value of 31.45 +/- 4.98 microM and a Vmax value of 30.90 +/- 1.18 pmol N-acetyltryptamine/min/mg protein for tryptamine, and a Km value of 28.72 +/- 7.50 microM and a Vmax value of 25.90 +/- 1.50 pmol N-acetyltryptamine/min/mg protein for acetyl CoA. The present method is simple, allows the determination of NAT activity from a variety of enzyme sources, has application to pharmacological studies of NAT regulation in tissue cultures, and provides an alternative to current radioenzymatic assays.

摘要

牛松果体及其他大鼠组织中血清素 - N - 乙酰基转移酶（NAT）活性的测定是基于通过高效液相色谱荧光检测法对由色胺和乙酰辅酶A形成的N - 乙酰色胺进行分离和检测。在牛松果体中，该酶对色胺的Km值为31.45±4.98微摩尔，Vmax值为30.90±1.18皮摩尔N - 乙酰色胺/分钟/毫克蛋白质；对乙酰辅酶A的Km值为28.72±7.50微摩尔，Vmax值为25.90±1.50皮摩尔N - 乙酰色胺/分钟/毫克蛋白质。本方法简单，可用于测定多种酶源的NAT活性，适用于组织培养中NAT调节的药理学研究，并为当前的放射酶法提供了一种替代方法。