Glass N L, Kosuge T
J Bacteriol. 1986 May;166(2):598-603. doi: 10.1128/jb.166.2.598-603.1986.
The phytopathogen Pseudomonas syringae subsp. savastanoi incites the production of galls on olive and oleander plants. Gall formation is dependent on bacterial production of the phytohormone indoleacetic acid (IAA). The genetic determinants for IAA synthesis are located on a plasmid (pIAA) and are organized in an operon in oleander strains of the bacterium. P. syringae subsp. savastanoi further converts IAA to an amino acid conjugate, 3-indole-acetyl-epsilon-L-lysine (IAA-lysine). The gene for IAA-lysine synthetase (iaaL) was found on the IAA plasmid by screening pIAA deletion mutants for the ability to convert IAA to IAA-lysine. The iaaL locus was then cloned in the vector pUC8 from a bank of P. syringae subsp. savastanoi EW2009 plasmid DNA to construct recombinant plasmid pLG87. The specific activity of IAA-lysine synthetase in Escherichia coli transformed with pLG87 was 47 times higher than that of the enzyme extract from P. syringae subsp. savastanoi. The direction of transcription of the iaaL gene was determined to be opposite to that of the IAA operon. The location of the iaaL gene on pIAA1 was mapped by Tn5 insertion mutagenesis to a 2.5-kilobase-pair fragment 2 kilobase pairs from the IAA operon.
植物病原菌丁香假单胞菌 savastanoi 亚种可在橄榄和夹竹桃植株上诱发瘿瘤形成。瘿瘤形成依赖于细菌产生的植物激素吲哚乙酸（IAA）。IAA 合成的遗传决定因素位于一个质粒（pIAA）上，并且在该细菌的夹竹桃菌株中以操纵子形式组织排列。丁香假单胞菌 savastanoi 亚种进一步将 IAA 转化为一种氨基酸共轭物，即 3 - 吲哚 - 乙酰 - ε - L - 赖氨酸（IAA - 赖氨酸）。通过筛选 pIAA 缺失突变体转化 IAA 为 IAA - 赖氨酸的能力，在 IAA 质粒上发现了 IAA - 赖氨酸合成酶（iaaL）基因。然后从丁香假单胞菌 savastanoi EW2009 质粒 DNA 文库中，将 iaaL 基因座克隆到载体 pUC8 中，构建重组质粒 pLG87。用 pLG87 转化的大肠杆菌中 IAA - 赖氨酸合成酶的比活性比丁香假单胞菌 savastanoi 亚种的酶提取物高 47 倍。已确定 iaaL 基因的转录方向与 IAA 操纵子的转录方向相反。通过 Tn5 插入诱变将 pIAA1 上的 iaaL 基因定位到距 IAA 操纵子 2 千碱基对的一个 2.5 千碱基对的片段上。
