The role of oxidoreductases in determining the function of the neisserial lipid A phosphoethanolamine transferase required for resistance to polymyxin.
The role of oxidoreductases in determining the function of the neisserial lipid A phosphoethanolamine transferase required for resistance to polymyxin.
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DOI：
10.1371/journal.pone.0106513
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发表时间：
2014
期刊：
影响因子：
3.7
通讯作者：
Kahler CM
中科院分区：
文献类型：
--
作者：
Piek S;Wang Z;Ganguly J;Lakey AM;Bartley SN;Mowlaboccus S;Anandan A;Stubbs KA;Scanlon MJ;Vrielink A;Azadi P;Carlson RW;Kahler CM
奈瑟氏菌属（Neisseria spp.）中脂寡糖（LOS）磷酸乙醇胺（PEA）转移酶（LptA）对LOS脂质A头基的修饰。是多粘菌素耐药性的核心。LptA的球状结构域的结构表明，该蛋白具有五个二硫键，表明它是细菌周质中蛋白质氧化途径的潜在底物。当奈瑟球菌LptA在大肠杆菌中表达的氧化还原酶，EcDsbA的存在下，多粘菌素耐药性增加了30倍。LptA修饰E. coli lipid A头部基团。在缺乏EcDsbA的情况下，LptA在E.杆菌奈瑟菌属表达三种氧化还原酶，DsbA 1，DsbA 2和DsbA 3，其中每一种似乎都向不同的靶标提供二硫键。N.脑膜炎球菌增强了对多粘菌素的敏感性，而组合突变体显示出对多粘菌素的敏感性的加性增加，这表明氧化还原酶是导致多粘菌素抗性的多种途径所必需的。寻找多粘菌素敏感性、LptA稳定性或活性与每种奈瑟球菌氧化还原酶存在之间的相关性。只有脑膜炎球菌突变体缺乏DsbA 3有一个可测量的减少PEA装饰的脂质A头基的量，这意味着LptA的稳定性支持的DsbA 3的存在下，但不需要DsbA 1/2，即使这些氧化还原酶可以氧化的蛋白质。这是第一个迹象表明，DsbA 3作为一个氧化还原酶在体内，多个氧化还原酶可能参与氧化的一个目标，在N。脑膜炎总之，LptA通过蛋白质内的二硫键稳定。当奈瑟球菌LptA在大肠杆菌中表达时，这种效果更加明显。coli中比N.可能反映了奈瑟球菌周质中的其他因子在LptA稳定性中起作用。
The decoration of the lipid A headgroups of the lipooligosaccharide (LOS) by the LOS phosphoethanolamine (PEA) transferase (LptA) in Neisseria spp. is central for resistance to polymyxin. The structure of the globular domain of LptA shows that the protein has five disulphide bonds, indicating that it is a potential substrate of the protein oxidation pathway in the bacterial periplasm. When neisserial LptA was expressed in Escherichia coli in the presence of the oxidoreductase, EcDsbA, polymyxin resistance increased 30-fold. LptA decorated one position of the E. coli lipid A headgroups with PEA. In the absence of the EcDsbA, LptA was degraded in E. coli. Neisseria spp. express three oxidoreductases, DsbA1, DsbA2 and DsbA3, each of which appear to donate disulphide bonds to different targets. Inactivation of each oxidoreductase in N. meningitidis enhanced sensitivity to polymyxin with combinatorial mutants displaying an additive increase in sensitivity to polymyxin, indicating that the oxidoreductases were required for multiple pathways leading to polymyxin resistance. Correlates were sought between polymyxin sensitivity, LptA stability or activity and the presence of each of the neisserial oxidoreductases. Only meningococcal mutants lacking DsbA3 had a measurable decrease in the amount of PEA decoration on lipid A headgroups implying that LptA stability was supported by the presence of DsbA3 but did not require DsbA1/2 even though these oxidoreductases could oxidise the protein. This is the first indication that DsbA3 acts as an oxidoreductase in vivo and that multiple oxidoreductases may be involved in oxidising the one target in N. meningitidis. In conclusion, LptA is stabilised by disulphide bonds within the protein. This effect was more pronounced when neisserial LptA was expressed in E. coli than in N. meningitidis and may reflect that other factors in the neisserial periplasm have a role in LptA stability.
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