Stalker D M, Hiatt W R, Comai L
J Biol Chem. 1985 Apr 25;260(8):4724-8.
The enzyme 5-enolpyruvylshikimate-3-phosphate synthase (EC 2.5.1.19), encoded by the aroA locus, is a target site of glyphosate inhibition in bacteria. A glyphosate-resistant aroA allele has been cloned in Escherichia coli from a mutagenized strain of Salmonella typhimurium. Subcloning of this mutant aroA allele shows the gene to reside on a 1.3-kilobase segment of S. typhimurium DNA. Nucleotide sequence analysis of this mutant gene indicates a protein-coding region 427 amino acids in length. Comparison of the mutant and wild type aroA gene sequences reveals a single base pair change resulting in a Pro to Ser amino acid substitution at the 101st codon of the protein. A hybrid gene fusion between mutant and wild type aroA gene sequences was constructed. 5-Enolpyruvylshikimate-3-phosphate synthase was prepared from E. coli cells harboring this construct. The glyphosate-resistant phenotype is shown to be associated with the single amino acid substitution described above.
由aroA基因座编码的5-烯醇丙酮酸莽草酸-3-磷酸合酶（EC 2.5.1.19）是细菌中草甘膦抑制的靶位点。已从鼠伤寒沙门氏菌的诱变菌株中克隆出抗草甘膦的aroA等位基因到大肠杆菌中。该突变aroA等位基因的亚克隆表明该基因位于鼠伤寒沙门氏菌DNA的1.3千碱基片段上。对该突变基因的核苷酸序列分析表明其蛋白质编码区长度为427个氨基酸。突变型和野生型aroA基因序列的比较揭示了一个单碱基对的变化，导致蛋白质第101个密码子处的脯氨酸被丝氨酸取代。构建了突变型和野生型aroA基因序列之间的杂交基因融合体。从携带该构建体的大肠杆菌细胞中制备了5-烯醇丙酮酸莽草酸-3-磷酸合酶。抗草甘膦表型显示与上述单个氨基酸取代有关。
