Regulation of Sesquiterpene Biosynthesis in Plants

甾醇和倍半萜的积累与异尾鲨类生物合成途径中特定分支点酶、角鲨烯合成酶和倍半萜环化酶的活性水平变化有关。下一个显而易见的问题是，这些酶的活性是否受到酶蛋白的绝对数量的调节。这两种酶都要纯化，每一种酶的抗体探针都要生成，每一种蛋白的绝对数量都要测定。用常规生化方法分离烟草细胞，测定两种途径共有的酶活性（3-羟基-3-甲基戊二酰辅酶A还原酶）和两种途径特有的酶活性（角鲨烯合成酶和倍半萜环化酶）的亚细胞分布。测量放射性甲戊酸盐、焦磷酸异戊烯基和焦磷酸法尼酯在体内与甾醇和倍半萜类化合物结合率的实验，以及随后的体外酶分析，将确定其他受调节的酶活性，从而可能限制酶活性。植物类异戊二烯包括一系列具有重要生理意义的化合物。植物生长激素)，以及农业(如。风味和香气成分)和药物(即。口服避孕药)的兴趣。我们对植物如何调节这些化合物的产生的理解仍然不完整。本研究旨在研究调节两类类异戊二烯、甾醇和倍半萜类生物合成的3种可能机制。这类信息应为未来农业和工业用植物的遗传和生物技术改良提供额外的战略

Accumulation of sterols and sesquiterpenoids have been correlated with changes in the activity level of particular branch point enzymes of the isopernoid biosynthetic pathway, squalene synthetase and sesquiterpene cyclase. The next obvious question is whether these enzyme activities are regulated by the absolute amounts of the enzyme proteins. Both enzymes are to be purified, antibody probes for each generated, and the absolute amount of each protein determined. The subcellular distribution of an enzyme activity common to both pathways, 3-hydroxy-3-methylglutaryl coenzyme A reductase, and those activities unique to each, squalene synthetase and sesquiterpene cyclase, will be determined in tobacco cells fractionated by conventional biochemical methods. Experiments measuring the in vivo incorporation rate of radioactive mevalonate, isopentenyl pyrophosphate, and farnesyl pyrophosphate into sterols and sesquiterpenoids, and subsequent in vitro enzyme assays will identify other regulated, and thus potentially limiting, enzyme activities.%%% Plant isoprenoids comprise a wide range of compounds of physiological importance (ie. plant growth hormones), as well as of agricultural (ie. flavor and aroma constituents) and pharmaceutical (ie. oral contraceptives) interest. Our understanding of how plants regulate the production of these compounds is still incomplete. The proposed work is to examine 3 putative mechanisms regulating the biosynthesis of two classes of isoprenoids, sterols and sesquiterpenoids. Information of this type should provide additional strategies for the future genetic and biotechnological improvement of plants for agriculture and industry.***//