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Studies on Transglutaminase Functioning as a Biomodulator Responsive to Food Environment

转谷氨酰胺酶作为响应食品环境的生物调节剂的研究

基本信息

批准号：
09660138
负责人：
IKURA Koji
金额：
$ 2.11万
依托单位：
Kyoto Institute of Technology
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (C)
财政年份：
1997
资助国家：
日本
起止时间：
1997 至 1998
项目状态：
已结题

项目摘要

This study was done to understand the relationship between the food environment and the functions of protein cross-linking enzyme transglutaminase being involved in various biological phenomena. ofmammals. 1. To isolate the amine acceptor protein substrates of transglutaminase in mammalian livers, a biotin-labeled primary amine substrate of transglutaminase, biotincadaverine, was used forbiotin labeling of proteins in the liver extracts by endogenous transglutaminase activity. The biotin-labeled proteins were isolated and recovered by biotin-avidin-affinity chromatography The obtainedproteins were separated by SDS-PAGE and their amino-terminal sequences were determined. The 38-kDa protein from rat liver was identified as a subunit of glyceraldehyde-3-phosphate dehydragenase, and the 28 kDa protein from guinea pig liver was identified as a subunit of glutathione S-transferase (classtheta). The 44 kDa proteins from mouse, rat, and guinea pig livers were identified as betaine-homocysteine S-methyltransferase. Both the glyceraldehyde-3-phosphate dehydrogenase from rabbitmuscle and glutathione S-transferase (class pi) from human placenta also could be amine acceptors in the amine incorporation catalyzed by guinea pig liver transglutaminase. These results suggested that these identified enzyme proteins can be modified post-translationally by cellular transglutaminase. 2. lnsulin, which lowers the blood glucose level and is invoked in the regulations of cell growth and differentiation, was tested for its signaling effect on the transglutaminase activity of human hepatoblastoma HepG2 cells. Insulin did not affect significantly the transglutaminase activity of HepG2 cells cultured in both media with and without dexamethasone. Transglutaminase activity of HepG2 cells also did not respond to the treatments of insulin-like growth factors I and II, and glucagon.

本研究旨在了解蛋白质交联酶转谷氨酰胺酶参与各种生物现象的功能与食物环境之间的关系。关于哺乳动物的。1.为了分离哺乳动物肝脏转谷氨酰胺酶的胺受体蛋白底物，将生物素标记的转谷氨酰胺酶伯胺底物Biotincadverine用于通过内源性转谷氨酰胺酶活性标记肝脏提取液中的蛋白质。用生物素-亲和素亲和层析法分离回收生物素标记的蛋白质，用十二烷基硫酸钠-聚丙烯酰胺凝胶电泳法分离蛋白质并测定其氨基末端序列。大鼠肝脏38 kDa蛋白为3-磷酸甘油醛脱氢酶亚基，豚鼠肝脏28 kDa蛋白为谷胱甘肽S转移酶亚基(Classtheta)。小鼠、大鼠和豚鼠肝脏的44 kDa蛋白质被鉴定为甜菜碱-同型半胱氨酸S-甲基转移酶。兔肌甘油醛-3-磷酸脱氢酶和人胎盘谷胱甘肽-S转移酶(pI类)在豚鼠肝脏谷氨酰胺转氨酶催化的胺掺入中也是胺受体。这些结果表明，这些已鉴定的酶蛋白可以被细胞转谷氨酰胺酶翻译后修饰。2.检测胰岛素对人肝母细胞瘤细胞株HepG2细胞谷氨酰胺转氨酶活性的信号转导作用。胰岛素对地塞米松和不加地塞米松培养的HepG2细胞谷氨酰胺转氨酶活性均无显著影响。细胞转谷氨酰胺酶活性对胰岛素样生长因子I和II以及胰高血糖素的处理也没有反应。