摘要
糖基供体通常以活性核糖的形式存在于细胞内,wbpX、wbpY以及wbpZ(糖基转移酶)等基因负责转运细胞内的活性糖基分子。糖基转移酶通过运送dTDP-鼠李糖(脂多糖合成前体之一)将鼠李糖基由细胞内转至胞外脂多糖O-抗原B-band部分。同时,参与鼠李糖脂合成的RhlB和RhlC(鼠李糖基转移酶)将dTDP-鼠李糖中的鼠李糖连接至3-羟基脂肪酸分别合成单、双鼠李糖脂。假单胞菌ZS1是本实验室在前期工作中从油污中分离出的1株高产鼠李糖脂菌株,本实验拟探究阻断脂多糖O-抗原B-band中鼠李糖基的添加对假单胞菌ZS1鼠李糖脂合成的影响。为此,构建了假单胞菌ZS1的wbpX、wbpY以及wbpZ 3个独立缺失突变菌株。结果表明,wbpX和wbpZ突变株的鼠李糖脂合成量均有增加,其中wbpZ的突变株合成量最高,较原菌株提高50%。SDS-PAGE分析结果表明,wbpZ缺失突变菌株的脂多糖O-抗原量显著降低;而wbpX和wbpY缺失突变菌株脂多糖含量变化不显著。
Glycosyl donors usually exist in cells in the form of active ribose,glycosyl transferases genes such as wbpX,wbpY and wbpZ are responsible for transporting active glycosyl molecules in cells.Glycosyltransferase transfers rhamnosyl from intracellular to extracellular lipopolysaccharide O-antigen B-band by transporting dTDP-rhamnose.Lipopolysaccharide is an important component of the cell wall of Gram-negative bacteria.Meanwhile,RhlB and RhlC(rhamnosyl transferases)involved in rhamnolipid synthesis link rhamnose in dTDP-rhamnose to 3-hydroxy fatty acids to synthesize mono-and di-rhamnolipids,respectively.Pseudomonas aeruginosa ZS1 is a high-yielding rhamnolipid strain isolated from oil stains in our laboratory in the previous work.This experiment intends to explore the effect of blocking the addition of rhamnosyl in the lipopolysaccharide O-antigen B-band on the synthesis of rhamnolipid in P.aeruginosa ZS1.To this end,we constructed three independent deletion mutants of wbpX,wbpY and wbpZ of P.aeruginosa ZS1.The results showed that both the wbpX and wbpZ mutants had increased rhamnolipid synthesis,and the wbpZ mutant had the highest synthesis ability,which was 50%higher than the original strain.The results of SDS-PAGE analysis showed that the content of lipopolysaccharide O-antigen in wbpZ deletion mutant strains decreased significantly,while the lipopolysaccharide content of wbpX and wbpY deletion mutant strains did not change significantly.
作者
张增平
黄小佩
马昀灏
杨支力
刘建华
ZHANG Zengping;HUANG Xiaopei;MA Yunhao(Pisa Mavine Graduate School of Zhejiang Ocean University,Zhoushan 316022,China)
出处
《浙江海洋大学学报（自然科学版）》
CAS
2023年第5期439-446,共8页
Journal of Zhejiang Ocean University：Natural Science
基金
国家自然科学基金面上项目(31571392)。
关键词
铜绿假单胞菌
鼠李糖脂
脂多糖合成
基因敲除
Pseudomonas aeruginosa
rhamnolipid
biosynthesis of LPS
gene knockout
