化学
戊二醛
酶
基质(水族馆)
膜
固定化酶
色谱法
共价键
酶分析
生物化学
有机化学
生物
生态学
作者
Nektaria Markoglou,Irving W. Wainer
出处
期刊:Journal of Biochemical and Biophysical Methods
[Elsevier]
日期:2001-03-01
卷期号:48 (1): 61-75
被引量:16
标识
DOI:10.1016/s0165-022x(00)00142-1
摘要
Dopamine beta-hydroxylase (DBH) catalyzes the beta-hydroxylation of dopamine to norepinephrine. The enzyme in chromaffin granules occurs in a soluble form and a form confined to the surrounding membrane. DBH was noncovalently immobilized in the hydrophobic interface of an immobilized artificial membrane (IAM) liquid chromatographic stationary phase and the resulting DBH-IAM stationary phase was enzymatically active and was shown to mimic the membrane-bound form of the enzyme. DBH was also covalently immobilized onto a silica-based support containing, glutaraldehyde-P (Glut-P). The resulting DBH-Glut-P interphase was also enzymatically active, reproducible and shown to display characteristics of the solubilized enzyme. The results demonstrate that the different immobilization methods utilized for the enzyme can be used to quantitatively and qualitatively determine the enzyme kinetic constants associated with enzyme/substrate and enzyme/inhibitor interactions for the two distinct forms of the enzyme. These new entities can be used in basic biochemical studies as well as in high throughput screening of substances for DBH substrate/inhibitor properties.
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