[目的]克隆佛手瓜鲨烯合酶（ SQS）基因，并基于克隆的SQS序列分析该酶的分子结构，为佛手瓜生物活性成分的利用提供理论依据。[方法]根据SQS基因5′末端保守序列和3′race方法扩增的佛手瓜SQS基因3′末端序列设计佛手瓜SQS基因克隆引物。扩增的佛手瓜SQS基因片段插入pGEM－T Easy Vector后经DNA测序。应用生物信息学方法分析克隆序列的开放阅读框架（ ORF）、酶蛋白的疏水性／亲水性和蛋白质磷酸化位点，预测酶蛋白的二级结构、结构域、三级结构和跨膜区。以布朗葡萄藻为outgroup，应用MEGA 5．0进行UPGMA算法构建系统树，并进行1000次的Bootstrap 测试。[结果]佛手瓜SQS由417氨基酸残基构成，等电点为6．983。构象预测提示该酶二级结构以α螺旋为主，是一种只有三级结构的单体酶，其活性中心是主要由几个α螺旋围绕形成的穴状结构，酶蛋白肽链中具有1个跨膜区。结构域分析表明SQS属于异戊二烯合酶家族。磷酸化位点分析显示该酶共有17个磷酸化位点，其中，S48位于与酶活性中心相关模体中，而S196为陆生植物SQS基因的正选择位点。以佛手瓜SQS基因ORF序列构建系统发生树得到的系统发生分类结果与形态学分类结果一致。[结论]佛手瓜SQS是由417氨基酸残基构成的单体酶，具有1个跨膜区和17个磷酸化位点，其中，S48和S196可能是佛手瓜SQS酶活性调节的关键性磷酸化位点。%Objective] To clone the gene of squalene synthase ( SQS) from Sechium edule Swartz and characterize the molecular structure of the enzyme, and thus provide the groundwork for exploiting the bioactive components of Sechium edule Swartz.[Method ] Primers for cloning of the SQS gene from Sechium edule Swartz were designed according to the 5′end conservative sequence of SQS genes and the 3′end sequence of the SQS gene from Sechium edule Swartz obtained by 3′race amplification.The amplified fragment of the SQS gene from Sechiume dule Swartz was inserted into the pGEM-T Easy Vector and verified by DNA sequencing.The analyses of the open reading frame (ORF ) of the Sechiu m eduleSwartz SQS gene sequence cloned, the hydrophobicity/hydrophily and the phosphorylation sites of the zymoprotein, as well as the predic-tions of the secondary structure, domain, tertiary structure and transmembrane region of the enzyme were conducted using bioinformatics meth-ods.The phylogenetic tress was constructed by performing the UPGMA of MEGA 5.0 and carrying out 1000 times of Bootstrap test, using bot-ryococcus braunii as outgroup.[Result] Sechium edule Swartz SQS is constituted of 417 amino acid residues with an isoelectric point of 6.983. Conformation prediction analyses suggested that αhelix is the main kind of the secondary structure in Sechium edule Swartz SQS zymoprotein, and the SQS is a monomeric enzyme with a cave-like activity center formed by αhelixes.Domain analysis showed that the enzyme molecule possesses a transmembrane region and suggested that the SQS is belonged to the isoprenoid biosynthase superfamily .And the analysis of phos-phorylation site indicated that there are 17 phosphorylation sites in the enzyme protein, among which the S48 locates in the activity center-relat-ed motif and the S196 is a positive selection site of the SQSgene .The result of phylogenetic classification based on the phylogenetic tress con-structed with the ORF sequence of the SQS gene was in accordance with that of morphologic classification.[Conclusion] Sechium edule Swartz SQS is a monomeric enzyme constituted of 417 amino acid residues and possesses a transmembrane region and 17 phosphorylation sites, among which the S48 and S196 may be the critical sites for the regulation of the SQS activity.
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