活动单胞菌和日本慢生根瘤菌合成类藿烷脂保守基因的克隆[j]

Michael Perzl , Ina G. Reipen , Susanne Schmitz , Karl Poralla , Hermann Sahm , Georg A. Sprenger , Elmar L. Kannenberg
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引用次数: 49

摘要

角鲨烯-藿烯环化酶(SHC)是唯一一种在遗传水平上被表征的参与类藿烷脂生物合成的酶。为了研究是否有其他参与藿烷类生物合成的基因与shc基因聚集在一起,我们克隆并分析了来自活动单胞菌和日本慢生根瘤菌的shc基因上游的核苷酸序列。在Z. mobilis中,检测到5个与shc基因紧密排列的开放阅读框(orf,命名为hpnA-E)。在B. japonicum中发现了3个类似排列的orf(对应于Z. mobilis的hpnC-E)。推导出的hpnC-E氨基酸序列在两种细菌中具有显著的相似性(58 ~ 62%)。与其他萜类生物合成途径(类胡萝卜素和类固醇生物合成)的酶相似,表明这些orf编码参与藿烷类及其中间体生物合成的蛋白质。在大肠杆菌中表达日本血吸虫的hpnC和Z. mobilis的hpnC导致了藿烷类前体角鲨烯的形成。这表明hpnC编码角鲨烯合成酶。另外两个orf (hpnA和hpnB)与参与糖转移和修饰的酶相似,表明它们可能编码参与藿烷类化合物含糖侧链复杂生物合成的酶。
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Cloning of conserved genes from Zymomonas mobilis and Bradyrhizobium japonicum that function in the biosynthesis of hopanoid lipids1

The squalene-hopene cyclase (SHC) is the only enzyme involved in the biosynthesis of hopanoid lipids that has been characterized on the genetic level. To investigate if additional genes involved in hopanoid biosynthesis are clustered with the shc gene, we cloned and analyzed the nucleotide sequences located immediately upstream of the shc genes from Zymomonas mobilis and Bradyrhizobium japonicum. In Z. mobilis, five open reading frames (ORFs, designated as hpnA–E) were detected in a close arrangement with the shc gene. In B. japonicum, three similarly arranged ORFs (corresponding to hpnC–E from Z. mobilis) were found. The deduced amino acid sequences of hpnC–E showed significant similarity (58–62%) in both bacteria. Similarities to enzymes of other terpenoid biosynthesis pathways (carotenoid and steroid biosynthesis) suggest that these ORFs encode proteins involved in the biosynthesis of hopanoids and their intermediates. Expression of hpnC to hpnE from Z. mobilis as well as expression of hpnC from B. japonicum in Escherichia coli led to the formation of the hopanoid precursor squalene. This indicates that hpnC encodes a squalene synthase. The two additional ORFs (hpnA and hpnB) in Z. mobilis showed similarities to enzymes involved in the transfer and modification of sugars, indicating that they may code for enzymes involved in the biosynthesis of the complex, sugar-containing side chains of hopanoids.

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