Menaquinone-specific prenyl reductase from the hyperthermophilic archaeon Archaeoglobus fulgidus
Menaquinone-specific prenyl reductase from the hyperthermophilic archaeon Archaeoglobus fulgidus
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DOI：
10.1128/jb.187.6.1937-1944.2005
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发表时间：
2005-03-01
影响因子：
3.2
通讯作者：
Nishino, T
中科院分区：
文献类型：
--
作者：
Hemmi, H;Takahashi, Y;Nishino, T
从嗜高温古菌Archaeoglobus fuldgidus中分离出4个编码植物香叶基香叶基还原酶同源物的基因，该古菌产生具有完全饱和的七异戊二烯基侧链的甲基萘醌，甲基萘醌-7（14 H）。同源物之一在大肠杆菌中的重组表达导致宿主细胞的醌概况的一个明显的变化，虽然同源物是最遥远的相关的香叶基香叶基还原酶。新化合物的洗脱时间依次长于普通醌类化合物。大肠杆菌，即，甲萘醌-8和泛醌-8，在反相柱上的高效液相色谱中。新化合物的结构分析表明，它们的分子量逐渐增加，相对于普通的醌在2 U的速率，但它们仍然包含醌头结构，强烈表明该化合物是醌与部分饱和的异戊二烯侧链。用连二亚硫酸盐作为还原剂的体外测定表明，异戊二烯基还原酶对甲基萘醌-7而不是泛醌-8和异戊二烯基二磷酸具有高度特异性。这种新的酶非共价结合黄素腺嘌呤二核苷酸，类似于香叶基香叶基还原酶，但不能利用NAD（P）H作为电子供体，不像植物同源物。
Four genes that encode the homologues of plant geranylgeranyl reductase were isolated from a hyperther-mophilic archaeon Archaeoglobus fuldgidus, which produces menaquinone with a fully saturated heptaprenyl side chain, menaquinone-7(14H). The recombinant expression of one of the homologues in Escherichia coli led to a distinct change in the quinone profile of the host cells, although the homologue is the most distantly related to the geranylgeranyl reductase. The new compounds found in the profile had successively longer elution times than those of ordinary quinones from E. coli, i.e., menaquinone-8 and ubiquinone-8, in high-performance liquid chromatography on a reversed-phase column. Structural analyses of the new compounds by electron impact-mass spectrometry indicated that their molecular masses progressively increase relative to the ordinary quinones at a rate of 2 U but that they still contain quinone head structures, strongly suggesting that the compounds are quinones with partially saturated prenyl side chains. In vitro assays with dithionite as the reducing agent showed that the prenyl reductase is highly specific for menaquinone-7, rather than ubiquinone-8 and prenyl diphosphates. This novel enzyme noncovalently binds flavin adenine dinucleotide, similar to geranylgeranyl reductase, but was not able to utilize NAD(P)H as the electron donor, unlike the plant homologue.