Clarification of the conversion mechanism of vitamin K analogues to menaquinone-4, a potent hormone-like substance, in the rat tissues.

阐明维生素 K 类似物在大鼠组织中向 Menaquinone-4（一种强效激素样物质）的转化机制。

基本信息

批准号：
11660119
负责人：
KOMAI Michio
金额：
$ 2.24万
依托单位：
Tohoku University
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (C)
财政年份：
1999
资助国家：
日本
起止时间：
1999 至 2000
项目状态：
已结题

项目摘要

Menaquinone-4 (MK-4) is one of naturally occurring form of vitmin K_2 (MK-n or K2) which is majorly synthesized in the microorganisms, but recently it has been shown that MK-4 is also synthesized from other vitamin K analogues in various tissues of rat and possibly in other mammalian tissues. According to recent investigations undertaken by us and other American and Dutch researchers, MK-4 abundantly distributes than other VK analogues to various tissues in the rats, such as testis, lung, brain, submandibullar gland, heart, spleen, liver, pancreas, and kidney. Recent results also showed that MK-4 is one of potent accelerator of bone formation in the rats, so MK-4 has been thought to be a potent hormone-like substance.We have newly established an MK-4 formation system in vitro using various tissues of the rats. Substrates for this reaction system are vitamin K_3 (menadione or VK3) or vitamin K_1 (phylloquinone or VK1) as a source for naphthoquinone ring, and geranylgeranyl-diphosphate a … More s a source for isoprenyl side-chain of MK-4, respectively. Supernatant fraction of 900 g of rat kidney homogenate with phophate buffer (pH 7.4) was used as crude enzyme solution. The present experiment undertaken for 2 years revealed that MK-4 production from VK3 was inhibited in a dose-related manner under the presence of warfarin that is an inhibitor for VKs epoxide and VKs reductases. This inhibition was recovered completely by an addition of reducing agent, di-thiothreitol (DTT). This lead us to consider that VK reductase participate in the conversion reaction from VK3 to MK-4, and that VK3-H_2 (reduced form of VK3) and MK-4-H_2 (reduced form of MK-4) might be involved in the reaction pathway as an intermediates. Purification trials of this key enzyme is now going on, and we are observing that the dissociation from the microsomal or mitochondrial membrane fraction made this enzyme much more unstable (less active).We have partly clarified the conversion mechanism of VKs into MK-4 much more clearly than before, and got useful information to purify this enzyme ([MK-4 synthase] as a tentative name). Further investigations are necessary to understand the physiological role of accumulated MK-4 in various tissues in the rat. Less

Menaquinone-4 (MK-4) 是维生素 K_2 (MK-n 或 K2) 的天然形式之一，主要在微生物中合成，但最近发现 MK-4 也可以由其他维生素 K 类似物合成根据我们以及其他美国和荷兰研究人员最近进行的研究，MK-4 在大鼠的各种组织中以及可能在其他哺乳动物组织中的分布比其他 VK 类似物更丰富。大鼠的睾丸、肺、脑、颌下腺、心脏、脾脏、肝脏、胰腺和肾脏等最近的研究结果也表明MK-4是大鼠骨形成的强效促进剂之一，因此MK-4已被广泛应用。被认为是一种有效的激素样物质。我们利用大鼠的各种组织在体外新建立了MK-4形成系统，该反应系统的底物是维生素K_3（甲萘醌或VK3）或维生素。 K_1（叶绿醌或 VK1）作为萘醌环的来源，香叶基香叶基二磷酸盐作为 MK-4 异戊二烯基侧链的来源，分别为 900 g 大鼠肾匀浆的上清液和磷酸盐缓冲液（pH 7.4）。本实验进行了 2 年，结果表明 VK3 的 MK-4 产生受到抑制。在作为 VK 环氧化物和 VK 还原酶抑制剂的华法林存在下，这种抑制作用通过添加还原剂二硫苏糖醇 (DTT) 完全恢复，这使我们认为 VK 还原酶参与了这一过程。从 VK3 到 MK-4 的转化反应，以及 VK3-H_2（VK3 的还原形式）和 MK-4-H_2（VK3 的还原形式） MK-4）可能作为中间体参与反应途径，目前正在进行这种关键酶的纯化试验，我们观察到从微粒体或线粒体膜部分解离使这种酶更加不稳定（活性较低）。 .我们已经比以前更清楚地部分阐明了VKs转化为MK-4的机制，并获得了纯化这种酶的有用信息（[MK-4合成酶]作为暂定名称）需要进一步研究以了解其生理作用。的大鼠各组织中MK-4的积累较少。