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甲羟戊酸激酶是植物中控制整个甲羟戊酸途径重要的限速酶,其代谢产物萜类物质是烟草芳香物质的主要来源。本研究采用同源克隆,利用RACE方法获得二倍体林烟草(Nicotiana sylvestris)甲羟戊酸激酶(NsyMK)的开放阅读框序列,对NsyMK序列进行生物信息学分析,预测蛋白理化性质、二级结构、三维结构并推测其结构与功能。结果表明,克隆获得的林烟草NsyMK的开放阅读框全长为1158bp,编码385个氨基酸,预测分子量41.4kDa、等电点为6.05,GenBank登录号KC871597。蛋白质结构分析显示NsyMK存在典型GHMP激酶和甲羟戊酸激酶保守结构域。生物信息学预测NsyMK蛋白不含跨膜区与信号肽。本研究成功克隆并分析了一个未报道的林烟草NsyMK的全长序列,为进一步阐明栽培烟草中萜类代谢途径奠定基础。
Mevalonate kinase is an important rate-limiting enzyme in plants that controls the entire mevalonate pathway and its metabolite, terpenoids, is a major source of tobacco aroma. In this study, homologous cloning was used to obtain the open reading frame of NsyMK from Nicotiana sylvestris by RACE method. Bioinformatics analysis of NsyMK sequence was carried out to predict the protein physical and chemical properties, Structure, three-dimensional structure and speculate on its structure and function. The results showed that the open reading frame of NsyMK was 1158bp in length and encoded a polypeptide of 385 amino acids. The predicted molecular weight was 41.4kDa and the isoelectric point was 6.05. GenBank accession number KC871597. Protein structure analysis showed that there are typical GHMP kinase and mevalonate kinase conserved domains in NsyMK. Bioinformatics prediction NsyMK protein does not contain the transmembrane region and signal peptide. In this study, we successfully cloned and analyzed a non-reported full-length sequence of NsyMK in N. tabaci, which laid the foundation for elucidating the terpenoid metabolic pathway in cultivated tobacco.