@article{ZaccheusBroekerLundborgetal.2012,
  author    = {Zaccheus, Mona V. and Br{\"o}ker, Nina Kristin and Lundborg, Magnus and Uetrecht, Charlotte and Barbirz, Stefanie and Widmalm, Goran},
  title     = {Structural studies of the O-antigen polysaccharide from Escherichia coli TD2158 having O18 serogroup specificity and aspects of its interaction with the tailspike endoglycosidase of the infecting bacteriophage HK620},
  series = {Carbohydrate research},
  volume    = {357},
  journal   = {Carbohydrate research},
  number    = {8},
  publisher = {Elsevier},
  address   = {Oxford},
  issn      = {0008-6215},
  doi       = {10.1016/j.carres.2012.05.022},
  pages     = {118 -- 125},
  year      = {2012},
  abstract  = {We have analyzed the O-antigen polysaccharide of the previously uncharacterized Escherichia coli strain TD2158 which is a host of bacteriophage HK620. This bacteriophage recognizes and cleaves the polysaccharide with its tailspike protein (TSP). The polysaccharide preparation as well as oligosaccharides obtained from HK620TSP endoglycosidase digests were analyzed with NMR spectroscopy. Additionally, sugar analysis was performed on the O-antigen polysaccharide and MALDI-TOF MS was used in oligosaccharide analysis. The present study revealed a heterogeneous polysaccharide with a hexasaccharide repeating unit of the following structure: alpha-D-Glcp-(1 -> 6) vertical bar vertical bar 2)-alpha-L-Rhap-(1 -> 6)-alpha-D-Glcp-(1 -> 4)-alpha-D-Galp-(1 -> 3)-alpha-D-GlcpNAc- (1 ->vertical bar beta-D-Glcp/beta-D-GlcpNAc-(1 -> 3) A repeating unit with a D-GlcNAc substitution of D-Gal has been described earlier as characteristic for serogroup O18A1. Accordingly, we termed repeating units with D-Glc substitution at D-Gal as O18A2. NMR analyses of the polysaccharide confirmed that O18A1- and O18A2-type repeats were present in a 1:1 ratio. However, HK620TSP preferentially bound the D-GlcNAc- substituted O18A1-type repeating units in its high affinity binding pocket with a dissociation constant of 140 mu M and disfavored the O18A2-type having a beta-D-Glcp-(1 -> 3)-linked group. As a result, in hexasaccharide preparations, O18A1 and O18A2 repeats were present in a 9: 1 ratio stressing the clear preference of O18A1- type repeats to be cleaved by HK620TSP.},
  language  = {en}
}
@article{BroekerGohlkeMuelleretal.2013,
  author    = {Br{\"o}ker, Nina Kristin and Gohlke, Ulrich and M{\"u}ller, J{\"u}rgen J. and Uetrecht, Charlotte and Heinemann, Udo and Seckler, Robert and Barbirz, Stefanie},
  title     = {Single amino acid exchange in bacteriophage HK620 tailspike protein results in thousand-fold increase of its oligosaccharide affinity},
  series = {Glycobiology},
  volume    = {23},
  journal   = {Glycobiology},
  number    = {1},
  publisher = {Oxford Univ. Press},
  address   = {Cary},
  issn      = {0959-6658},
  doi       = {10.1093/glycob/cws126},
  pages     = {59 -- 68},
  year      = {2013},
  abstract  = {Bacteriophage HK620 recognizes and cleaves the O-antigen polysaccharide of Escherichia coli serogroup O18A1 with its tailspike protein (TSP). HK620TSP binds hexasaccharide fragments with low affinity, but single amino acid exchanges generated a set of high-affinity mutants with submicromolar dissociation constants. Isothermal titration calorimetry showed that only small amounts of heat were released upon complex formation via a large number of direct and solvent-mediated hydrogen bonds between carbohydrate and protein. At room temperature, association was both enthalpy- and entropy-driven emphasizing major solvent rearrangements upon complex formation. Crystal structure analysis showed identical protein and sugar conformers in the TSP complexes regardless of their hexasaccharide affinity. Only in one case, a TSP mutant bound a different hexasaccharide conformer. The extended sugar binding site could be dissected in two regions: first, a hydrophobic pocket at the reducing end with minor affinity contributions. Access to this site could be blocked by a single aspartate to asparagine exchange without major loss in hexasaccharide affinity. Second, a region where the specific exchange of glutamate for glutamine created a site for an additional water molecule. Side-chain rearrangements upon sugar binding led to desolvation and additional hydrogen bonding which define this region of the binding site as the high-affinity scaffold.},
  language  = {en}
}