Elucidation of Reaction Mechanisms of Chain-elongation Enzymes in Isoprenoid Biosynthesis

1.对嗜热脂肪芽孢杆菌耐热的法尼基二磷酸合成酶进行了定点突变，并对其结构与活性的关系进行了研究。结果表明，保守基序FQXXDD中的Phe-220和Gln-221都是催化所必需的，Lys-47和Lys-183参与了异戊烯基二磷酸的结合。综上所述，这些发现有助于更好地理解异丙烯基转移酶反应的机制。克隆、测序并高效表达了嗜热杆菌七烯基二磷酸合成酶的两个组分的基因。编码甲基化转移酶的基因被发现位于七烯基二磷酸合成酶的两个基因之间，该基因负责菜醌生物合成的最后一步。实现了牛脑香叶基香叶基二磷酸合成酶的纯化，明确了其催化功能。通过研究甲氧丙戊酸在盐生细菌中的代谢，发现了一种新型的类异戊二烯衍生材料修饰蛋白质。在葡萄孢子菌中发现了法尼醇磷酸激酶，并对其性质进行了揭示。发现了一种新的代谢系统，可将多酚和脱氢多酚转化为它们的醛。

1. Site-directed mutations were introduced into thermostable farnesyl diphosphate synthase of Bacillus stearothermophilus, and structure-reactivity relation was investigated. As a result, it has been demonstrated that Phe-220 and Gln-221 in the conserved motif FQXXDD are both essential for catalysis and that Lys-47 and Lys-183 are involved in the binding of isopentenyl diphosphate. Coupled together, these findings have led to a better understanding of the mechanism of prenyltransferase reaction.2. The genes for the two components of B.stearothermophilus heptaprenyl diphosphate synthase were cloned, sequenced, and overexpressed. The gene encoding a methyltransferease responsible for the final step in menaquinone biosynthesis was found to be located between the two genes for heptaprenyl diphosphate synthase.3. The purification of geranylgeranyl diphosphate synthase from bovine brain was achived, and the catalytic function was clarified.4. A novel type of protein modification by isoprenoid derived materials was discovered by investigating the metabolism of mevalonate in halobacterium halobium.5. Farnesol phosphokinase was found in Botryococcus braunii and its properties was disclosed.6. A new metabolic system for the conversion of dolichols and dehydrodolichols into their aldehydes was discovered.