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DOI | 10.1006/abbi.2000.1731 |
Characterization of ecdysteroid 26-hydroxylase: An enzyme involved in molting hormone inactivation | |
Williams, DR; Fisher, MJ; Rees, HH | |
通讯作者 | Rees, HH |
来源期刊 | ARCHIVES OF BIOCHEMISTRY AND BIOPHYSICS
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ISSN | 0003-9861 |
出版年 | 2000 |
卷号 | 376期号:2页码:389-398 |
英文摘要 | Insect molting hormone (ecdysteroid) inactivation occurs by several routes, including 26-hydroxylation and further oxidation to the 26-oic acids. Thus, the ecdysteroid 26-hydroxylase is a critical enzyme involved in precise regulation of ecdysteroid titers during insect development. Administration of the ecdysteroid agonist, RH-5849 (1,2-dibenzoyl, 1-tert-butyl hydrazone), or 20-hydroxyecdysone to the tobacco hornworm, Manduca sexta, results in induction of ecdysteroid 26-hydroxylase activity in midgut mitochondria and microsomes. The biochemical and kinetic properties of the ecdysteroid 26-hydroxylase were investigated. The mitochondrial enzyme was found to have optimal activity at a pH of 7.5 in a Hepes or sodium phosphate buffer at 30-37 degrees C. The apparent K-m of the microsomal 26-hydroxylase for 20-hydroxyecdysone substrate was lower than that of the mitochondrial enzyme for either 20-hydroxyecdysone or ecdysone substrate. The V-max of the 26-hydroxylase in both subcellular fractions was slightly higher using 20-hydroxyecdysone as substrate compared to ecdysone, Demonstration that activity of the mitochondrial 26-hydroxylase was inhibited by incubation in a CO (or N-2) atmosphere, taken together with the requirement for reducing cofactor and the efficacy of the P450 inhibitors, ketoconazole and fenarimol, provided strong evidence that the hydroxylase is cytochrome P450-dependent. Indirect evidence suggested that the mitochondrial and microsomal ecdysteroid 26-hydroxylase(s) could exist in a less active dephosphorylated state or more active phosphorylated state. Using Escherichia coli alkaline phosphatase to remove covalently bound phosphate groups, the activity of the 26-hydroxylase was decreased and, conversely, activity was enhanced using a cAMP-dependent protein kinase with appropriate cofactors, In addition, the protein kinase was shown to reactivate the 26-hydroxylase activity in alkaline phosphatase-treated fractions. (C) 2000 academic Press. |
英文关键词 | ecdysteroid 26-hydroxylase cytochrome P450 Manduca sexta |
类型 | Article |
语种 | 英语 |
国家 | England |
收录类别 | SCI-E |
WOS记录号 | WOS:000086488900020 |
WOS关键词 | ECDYSONE 20-MONOOXYGENASE ACTIVITY ; MANDUCA-SEXTA ; CYTOCHROME-P-450 ENZYMES ; SCHISTOCERCA-GREGARIA ; REVERSIBLE ACTIVATION ; DESERT LOCUST ; PHOSPHORYLATION ; MITOCHONDRIAL ; 20-MONO-OXYGENASE ; BIOSYNTHESIS |
WOS类目 | Biochemistry & Molecular Biology ; Biophysics |
WOS研究方向 | Biochemistry & Molecular Biology ; Biophysics |
资源类型 | 期刊论文 |
条目标识符 | http://119.78.100.177/qdio/handle/2XILL650/138574 |
作者单位 | (1)Univ Liverpool, Sch Biol Sci, Cellular Regulat & Signaling Grp, Liverpool L69 7ZB, Merseyside, England |
推荐引用方式 GB/T 7714 | Williams, DR,Fisher, MJ,Rees, HH. Characterization of ecdysteroid 26-hydroxylase: An enzyme involved in molting hormone inactivation[J],2000,376(2):389-398. |
APA | Williams, DR,Fisher, MJ,&Rees, HH.(2000).Characterization of ecdysteroid 26-hydroxylase: An enzyme involved in molting hormone inactivation.ARCHIVES OF BIOCHEMISTRY AND BIOPHYSICS,376(2),389-398. |
MLA | Williams, DR,et al."Characterization of ecdysteroid 26-hydroxylase: An enzyme involved in molting hormone inactivation".ARCHIVES OF BIOCHEMISTRY AND BIOPHYSICS 376.2(2000):389-398. |
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