Escherichia coli O157:H7 Strain EDL933 Harbors Multiple Functional Prophage-Associated Genes Necessary for the Utilization of 5-N-Acetyl-9-O-Acetyl Neuraminic Acid as a Growth Substrate
Escherichia coli O157:H7 Strain EDL933 Harbors Multiple Functional Prophage-Associated Genes Necessary for the Utilization of 5-N-Acetyl-9-O-Acetyl Neuraminic Acid as a Growth Substrate
复制标题
DOI：
10.1128/aem.01671-16
复制
发表时间：
2016-10-01
影响因子：
4.4
通讯作者：
Schmidt, Herbert
中科院分区：
文献类型：
--
作者：
Saile, Nadja;Voigt, Anja;Schmidt, Herbert
肠出血性大肠杆菌(EHEC)O157：H7株EDL933在其基因组中含有多个与染色体nans基因高度同源的前噬菌体相关开放阅读框架(ORF)。后者是纳米CMS操纵子的一部分，该操纵子存在于大多数大肠杆菌菌株中，编码一种酯酶，负责5-N-乙酰基-9-O-乙酰神经氨酸(Neu5，9Ac(2))的单一脱乙酰基。虽然一个原噬菌体携带的ORF(Z1466)已经在先前的研究中被鉴定，但其他NANS同源ORF的功能尚不清楚。在本研究中，我们初步研究了EDL933的nans同源ORF。根据它们与染色体nans基因的同源性和它们在噬菌体基因组中的位置，我们将它们命名为nans-p，并将不同的nans-p等位基因从1到10连续编号。我们选择了两个等位基因nans-p2和nans-p4来生产重组蛋白，并对它们的酶活性进行了研究，发现它们的最适温度存在差异。此外，在以Neu5，9Ac(2)为碳源和添加不同重组nans-p蛋白的生长培养基中，这些酶在底物利用中的功能可以用E.coliC600 Delta nans突变体来证明。此外，菌株EDL933中所有nans-p等位基因的连续缺失的产生和随后的生长实验表明，Neu5，9Ac(2)的利用存在基因剂量效应。由于Neu5，9Ac(2)是人和动物肠道粘液的重要组成部分，而且在大肠中的营养可获得性有限，我们假设，在特定条件下，多个Neu5，9Ac(2)酯酶的存在为它们在大肠中提供了营养供应，即使特定的前驱体丢失了。
Enterohemorrhagic Escherichia coli (EHEC) O157:H7 strain EDL933 harbors multiple prophage-associated open reading frames (ORFs) in its genome which are highly homologous to the chromosomal nanS gene. The latter is part of the nanCMS operon, which is present in most E. coli strains and encodes an esterase which is responsible for the monodeacetylation of 5-N-acetyl-9-O-acetyl neuraminic acid (Neu5,9Ac(2)). Whereas one prophage-borne ORF (z1466) has been characterized in previous studies, the functions of the other nanS-homologous ORFs are unknown. In the current study, the nanS-homologous ORFs of EDL933 were initially studied in silico. Due to their homology to the chromosomal nanS gene and their location in prophage genomes, we designated them nanS-p and numbered the different nanS-p alleles consecutively from 1 to 10. The two alleles nanS-p2 and nanS-p4 were selected for production of recombinant proteins, their enzymatic activities were investigated, and differences in their temperature optima were found. Furthermore, a function of these enzymes in substrate utilization could be demonstrated using an E. coli C600 Delta nanS mutant in a growth medium with Neu5,9Ac(2) as the carbon source and supplementation with the different recombinant NanS-p proteins. Moreover, generation of sequential deletions of all nanS-p alleles in strain EDL933 and subsequent growth experiments demonstrated a gene dose effect on the utilization of Neu5,9Ac(2). Since Neu5,9Ac(2) is an important component of human and animal gut mucus and since the nutrient availability in the large intestine is limited, we hypothesize that the presence of multiple Neu5,9Ac(2) esterases provides them a nutrient supply under certain conditions in the large intestine, even if particular prophages are lost.