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@ARTICLE{Losasso:201291,
      author       = {Losasso, Valeria and Schiffer, Sonja and Barth, Stefan and
                      Carloni, Paolo},
      title        = {{D}esign of human granzyme {B} variants resistant to serpin
                      {B}9},
      journal      = {Proteins},
      volume       = {80},
      number       = {11},
      issn         = {0887-3585},
      address      = {New York, NY},
      publisher    = {Wiley-Liss},
      reportid     = {FZJ-2015-03596},
      pages        = {2514 - 2522},
      year         = {2012},
      abstract     = {Human granzyme B (hGB) is a serine protease involved in
                      immune-mediated apoptosis. Its cytotoxicity makes it
                      potentially applicable in cancer therapy. However, the
                      effectiveness of hGB can be hampered by the cytosolic
                      expression of a natural protein inhibitor, human Serpin B9
                      (hSB9). Here, we used computational approaches to identify
                      hGB mutations that can affect its binding to hSB9 without
                      significantly decreasing its catalytic efficiency.
                      Alanine-scanning calculations allowed us to identify
                      residues of hGB important for the interaction with hSB9.
                      Some variants were selected, and molecular dynamic
                      simulations on the mutated hGB in complex with hSB9 in
                      aqueous solution were carried out to investigate the effect
                      of these variants on the stability of the complex. The R28K,
                      R201A, and R201K mutants significantly destabilized the
                      interaction of the protein with hSB9. Consistently, all of
                      these variants also retained their activity in the presence
                      of the Serpin B9 inhibitor in subsequent in vitro assays of
                      wild-type and mutated hGB. In particular, the activity of
                      R201K hGB with and without Serpin B9 is very similar to that
                      of the wild-type protein. Hence, R201K hGB emerges as a
                      promising species for antitumoral therapy applications},
      cin          = {GRS / IAS-5},
      ddc          = {540},
      cid          = {I:(DE-Juel1)GRS-20100316 / I:(DE-Juel1)IAS-5-20120330},
      pnm          = {899 - ohne Topic (POF2-899)},
      pid          = {G:(DE-HGF)POF2-899},
      typ          = {PUB:(DE-HGF)16},
      UT           = {WOS:000309456500003},
      pubmed       = {pmid:22733450},
      doi          = {10.1002/prot.24133},
      url          = {https://juser.fz-juelich.de/record/201291},
}