论文部分内容阅读
烯脂酰ACP还原酶是细菌脂肪酸合成的关键酶之一.本研究通过生物信息学分析发现,野油菜黄单胞菌Xanthomonas campestris(Xcc)8004基因组中XC_0119(Xccfab V)注释为反-2-烯脂酰Co A还原酶基因.但其编码产物与铜绿假单胞菌的烯脂酰ACP还原酶Fab V具有较高的同源性,并含有相同的催化活性中心Tyr-(Xaa)8-Lys序列.用携带Xccfab V的质粒载体互补大肠杆菌fab I温度敏感突变株JP1111,转化子能在42℃生长,表明Xccfab V能遗传互补大肠杆菌fab I突变.体外重建脂肪酸合成反应表明,Xcc Fab V能催化不同链长的烯脂酰ACP还原为脂酰ACP,且催化活性不受三氯森抑制.遗传学研究表明,Xccfab V是必需基因,不能获得Xccfab V基因敲除突变株.将携带大肠杆菌fab I的外源质粒导入野生菌后,可敲除染色体上的fab V基因,获得的替换突变株生长特性和脂肪酸组成未发生显著变化,但替换突变株对三氯森敏感.上述结果证实,野油菜黄单胞菌fab V是必需基因,编码烯脂酰ACP还原酶,参与脂肪酸从头合成反应,且Fab V是Xcc对三氯森耐受的根本原因.
Enolase ACP reductase is one of the key enzymes in bacterial fatty acid synthesis.In this study, bioinformatics analysis showed that XC_0119 (Xccfab V) in Xanthomonas campestris (Xcc) 8004 genome was annotated as trans-2- However, the encoded product has high homology with the Fab V of the ACO reductase of Pseudomonas aeruginosa and contains the same catalytic active site Tyr- (Xaa) 8- Lys sequence.With the plasmid vector carrying Xccfab V, the temperature-sensitive E. coli fab I strain JP1111 was mutated and the transformants were able to grow at 42 ℃, indicating that Xccfab V was able to mutate the FabⅠ mutant of E. coli in vitro. The in vitro reconstitution of fatty acid showed that Xcc Fab V can catalyze the reduction of acyl acyl-ACP with different chain lengths to acyl-ACP, and its catalytic activity is not inhibited by triclosan.Genetic studies show that Xccfab V is an essential gene and can not obtain Xccfab V-gene knockout mutants. After the introduction of the wild-type plasmid of E. coli fab I into the wild-type strain, the fab V gene on the chromosome was knocked out and the growth characteristics and fatty acid composition of the obtained mutant were not significantly changed, but the mutant was sensitive to triclosan. certificate , Xanthomonas campestris fab V is essential gene encoding enoyl-ACP reductase, involved in de novo fatty acid synthesis, and Xcc Fab V is the root causes of Triclosan tolerated.