Department of Biotechnology, Panjab University, Chandigarh 160014, India.
Mol Biotechnol. 2013 Jul;54(3):944-53. doi: 10.1007/s12033-012-9645-1.
Saponins are important group of secondary metabolites known for their pharmacological properties. Chlorophytum borivilianum contains high amount of saponins and is thus, recognized as an important medicinal plant with aphrodisiac properties. Though the plant is well known for its pharmaceutical properties, there is meager information available about the genes and enzymes responsible for biosynthesis of saponins from this plant. Squalene synthase (SqS) is the key enzyme of saponin biosynthesis pathway and here, we report cloning and characterization of SqS gene from C. borivilianum. A full-length CbSqS cDNA consisting of 1,760 bp was cloned which contained an open reading frame (ORF) of 1,233 bp, encoding a protein of 411 amino acids. Analysis of deduced amino acid sequence of CbSqS predicted the presence of conserved isoprenoid family domain and catalytic sites. Phylogenetic analysis revealed that CbSqS is closer to Glycine max and monocotyledonous plants. 3D structure prediction using various programs showed CbSqS structure to be similar to SqS from other species. C-terminus truncated recombinant squalene synthase (TruncCbSqS) was expressed in E. coli M15 cells with optimum expression induced with 1 mM IPTG at 37 °C. The gene expression level was analyzed through semi-quantitative RT-PCR and was found to be higher in leaves as compared to the roots.
皂素是一类具有重要药理活性的次生代谢产物。宽叶吊兰含有大量的皂素，因此被认为是一种具有壮阳作用的重要药用植物。尽管该植物因其药用特性而广为人知，但关于其合成皂素的基因和酶的信息却很少。鲨烯合酶（SqS）是皂素生物合成途径的关键酶，在此，我们报道了从宽叶吊兰中克隆和鉴定 SqS 基因。克隆得到一个全长为 1760bp 的 CbSqS cDNA，其中包含一个 1233bp 的开放阅读框（ORF），编码一个 411 个氨基酸的蛋白质。对 CbSqS 推导的氨基酸序列分析预测存在保守的异戊二烯家族结构域和催化位点。系统发育分析表明 CbSqS 与 Glycine max 和单子叶植物更为接近。使用各种程序对 3D 结构进行预测表明，CbSqS 结构与其他物种的 SqS 相似。在大肠杆菌 M15 细胞中表达了 C 端截断的重组鲨烯合酶（TruncCbSqS），最佳诱导表达条件为 37°C、1mM IPTG。通过半定量 RT-PCR 分析基因表达水平，发现与根相比，叶中的基因表达水平更高。
