番茄醇酰基转移酶基因SlAAT1克隆、序列分析和原核表达

酯类物质是许多果实香气的主要成分。醇酰基转移酶(AATs)是酯类化合物合成的关键酶。本研究通过反转录PCR，从番茄的成熟果实中克隆了SlAAT1基因(GenBank登录号为JQ070977)，其编码一个含有442个氨基酸残基的蛋白，含有醇酰转移酶BAHD家族的H-x-x-x-D和DFGWG保守基序。系统进化分析表明，SlAAT1与苹果MpAAT1，山字草的BEBT及烟草Hsr201等聚在同一分支，进化关系较近。SDS-PAGE电泳分析表明，转化SlAAT1基因的大肠杆菌BL21(DE3)在22℃、0.8 mmol·L^-1 IPTG条件下可获得大量的可溶性目标蛋白。同时，纯化的SlAAT1大肠杆菌重组蛋白的体外酶活性分析表明了SlAAT1重组蛋白具有醇酰基转移酶活性，可能参与了酯类挥发性成分的合成。

Cloning,Sequence Analysis and Prokaryotic Expression of an Alcohol Acyltransferase(AAT) Gene in Tomato(Solanum lycopersicum)

Volatile esters are important aromatic compounds accumulated in many ripe fruits. Alcohol acyltransferase (AAT) plays a key role in the formation of volatile esters. In the present study, a full-length cDNA sequence homologous with other plant AAT genes was isolated by RT-PCR from tomato ripe fruit, and named as SlAAT1(GenBank Accession No.JN398667). Results of sequence analyses showed SlAAT1 encodes a 442-amino acid protein, which exhibits conserved features of the BAHD family of acetyl transferase, such as HxxxD and DFGWG motifs. Phylogenetic analysis revealed that SlAAT1 is very closely related to the apple MpAAT1, Clarkia breweri BEBT, and tobacco Hsr201. The recombinant plasmid was transformed into E. coil BL21 (DE3) to express the SlAAT1-His6 protein in the optimized condition (22℃, 0.8 mmol·L^-1 IPTG). The purified recombinant SlAAT1 protein was shown to have acetyltransferase activity, suggesting its role in volatile ester formation.