Roles of predicted glycosyltransferases in the biosynthesis of the Rhizobium etli CE3 O antigen.
Roles of predicted glycosyltransferases in the biosynthesis of the Rhizobium etli CE3 O antigen.
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预测的糖基转移酶在根瘤菌 CE3 O 抗原生物合成中的作用。
DOI：
10.1128/jb.02080-12
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发表时间：
2013
影响因子：
3.2
通讯作者：
Noel,KDale
中科院分区：
文献类型：
--
作者：
Ojeda,KristyleaJ;Simonds,Laurie;Noel,KDale
根瘤菌CE 3 O抗原是一种固定长度的杂聚物。其合成所需的遗传区域已经确定，并且核苷酸序列是已知的。O抗原的结构已经确定，但特定基因在合成这种结构中的作用相对不清楚。在已知的O抗原基因簇中，发现9个开放阅读框（ORF）含有保守的糖基转移酶结构域。对每个ORF进行突变，并分析所得突变体脂多糖（LPS）。Tricine SDS-PAGE显示O抗原的逐步截短，这与突变体LPS糖组成的差异和与O抗原特异性单克隆抗体的反应性一致。根据这些结果和目前的理论，O-抗原合成，推导出特定的作用，为九个糖基转移酶的每一个，和生物合成的R。Etli CE 3 O抗原。在该模型中，通过糖基转移酶WreU将N-乙酰基-奎诺沃胺-磷酸（QuiNAc-P）添加至巴曲雷诺-磷酸来启动O-抗原生物合成。糖基转移酶WreG、WreE、WreS和WreT将分别作用一次以连接甘露糖、岩藻糖、第二岩藻糖和3-O-甲基-6-脱氧塔罗糖（3 OMe 6dTal）。然后，WreH将催化甲基葡萄糖醛酸（MeGlcA）的加成，以完成O-抗原重复单元的第一个实例。由岩藻糖、3 OMe 6dTal和MeGlcA组成的该单元的四个后续重复将通过由两种另外的糖基转移酶WreM和WreL与WreH一起沿着催化的反应循环来组装。最后，O抗原将通过糖基转移酶WreB催化的二-或三-O-甲基化岩藻糖的连接而加帽。
The Rhizobium etli CE3 O antigen is a fixed-length heteropolymer. The genetic regions required for its synthesis have been identified, and the nucleotide sequences are known. The structure of the O antigen has been determined, but the roles of specific genes in synthesizing this structure are relatively unclear. Within the known O-antigen genetic clusters of this strain, nine open reading frames (ORFs) were found to contain a conserved glycosyltransferase domain. Each ORF was mutated, and the resulting mutant lipopolysaccharide (LPS) was analyzed. Tricine SDS-PAGE revealed stepwise truncations of the O antigen that were consistent with differences in mutant LPS sugar compositions and reactivity with O-antigen-specific monoclonal antibodies. Based on these results and current theories of O-antigen synthesis, specific roles were deduced for each of the nine glycosyltransferases, and a model for biosynthesis of the R. etli CE3 O antigen was proposed. In this model, O-antigen biosynthesis is initiated with the addition ofN-acetyl-quinovosamine-phosphate (QuiNAc-P) to bactoprenol-phosphate by glycosyltransferase WreU. Glycosyltransferases WreG, WreE, WreS, and WreT would each act once to attach mannose, fucose, a second fucose, and 3-O-methyl-6-deoxytalose (3OMe6dTal), respectively. WreH would then catalyze the addition of methyl glucuronate (MeGlcA) to complete the first instance of the O-antigen repeat unit. Four subsequent repeats of this unit composed of fucose, 3OMe6dTal, and MeGlcA would be assembled by a cycle of reactions catalyzed by two additional glycosyltransferases, WreM and WreL, along with WreH. Finally, the O antigen would be capped by attachment of di- or tri-O-methylated fucose as catalyzed by glycosyltransferase WreB.