title: 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.
abstract: 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 (Neu5,9Ac2). 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 C600ΔnanS mutant in a growth medium with Neu5,9Ac2 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 Neu5,9Ac2 Since Neu5,9Ac2 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 Neu5,9Ac2 esterases provides them a nutrient supply under certain conditions in the large intestine, even if particular prophages are lost.
  
  
  
  In this study, a group of homologous prophage-borne nanS-p alleles and two of the corresponding enzymes of enterohemorrhagic E. coli (EHEC) O157:H7 strain EDL933 that may be important to provide alternative genes for substrate utilization were characterized.
authors.affiliation: Department of Food Microbiology and Hygiene, Institute of Food Science and Biotechnology, University of Hohenheim, Stuttgart, Germany.
authors.forename: Nadja
authors.initials: N
authors.lastname: Saile
authors.affiliation: Department of Food Microbiology and Hygiene, Institute of Food Science and Biotechnology, University of Hohenheim, Stuttgart, Germany.
authors.forename: Anja
authors.initials: A
authors.lastname: Voigt
authors.affiliation: Department of Food Microbiology and Hygiene, Institute of Food Science and Biotechnology, University of Hohenheim, Stuttgart, Germany.
authors.forename: Sarah
authors.initials: S
authors.lastname: Kessler
authors.affiliation: Department of Biotechnology and Enzyme Science, Institute of Food Science and Biotechnology, University of Hohenheim, Stuttgart, Germany.
authors.forename: Timo
authors.initials: T
authors.lastname: Stressler
authors.affiliation: Department of Health Sciences and Technology, Institute of Food, Nutrition and Health, ETH Zurich, Zurich, Switzerland.
authors.forename: Jochen
authors.identifier: 0000-0003-3410-2702
authors.initials: J
authors.lastname: Klumpp
authors.affiliation: Department of Biotechnology and Enzyme Science, Institute of Food Science and Biotechnology, University of Hohenheim, Stuttgart, Germany.
authors.forename: Lutz
authors.initials: L
authors.lastname: Fischer
authors.affiliation: Department of Food Microbiology and Hygiene, Institute of Food Science and Biotechnology, University of Hohenheim, Stuttgart, Germany herbert.schmidt@uni-hohenheim.de.
authors.forename: Herbert
authors.initials: H
authors.lastname: Schmidt
mesh_terms: D003198:Computer Simulation; D019453:Escherichia coli O157 / Q000235:genetics* / Q000254:growth & development / Q000378:metabolism; D029968:Escherichia coli Proteins / Q000235:genetics / Q000378:metabolism; D016366:Open Reading Frames; D039002:Prophages / Q000235:genetics* / Q000378:metabolism; D017384:Sequence Deletion; D012794:Sialic Acids / Q000378:metabolism*
chemical_list: D029968:Escherichia coli Proteins; D012794:Sialic Acids; C025676:9-O-acetyl-N-acetylneuraminic acid