@phdthesis{ReynaGonzalez2017,
  author    = {Reyna Gonz{\´a}lez, Emmanuel},
  title     = {Engineering of the microviridin post-translational modification enzymes for the production of synthetic protease inhibitors},
  url       = {http://nbn-resolving.de/urn:nbn:de:kobv:517-opus4-406979},
  school      = {Universit{\"a}t Potsdam},
  pages     = {XI, 91, CI},
  year      = {2017},
  abstract  = {Natural products and their derivatives have always been a source of drug leads. In particular, bacterial compounds have played an important role in drug development, for example in the field of antibiotics. A decrease in the discovery of novel leads from natural sources and the hope of finding new leads through the generation of large libraries of drug-like compounds by combinatorial chemistry aimed at specific molecular targets drove the pharmaceutical companies away from research on natural products. However, recent technological advances in genetics, bioinformatics and analytical chemistry have revived the interest in natural products. The ribosomally synthesized and post-translationally modified peptides (RiPPs) are a group of natural products generated by the action of post-translationally modifying enzymes on precursor peptides translated from mRNA by ribosomes. The great substrate promiscuity exhibited by many of the enzymes from RiPP biosynthetic pathways have led to the generation of hundreds of novel synthetic and semisynthetic variants, including variants carrying non-canonical amino acids (ncAAs). The microviridins are a family of RiPPs characterized by their atypical tricyclic structure composed of lactone and lactam rings, and their activity as serine protease inhibitors. The generalities of their biosynthetic pathway have already been described, however, the lack of information on details such as the protease responsible for cleaving off the leader peptide from the cyclic core peptide has impeded the fast and cheap production of novel microviridin variants. In the present work, knowledge on leader peptide activation of enzymes from other RiPP families has been extrapolated to the microviridin family, making it possible to bypass the need of a leader peptide. This feature allowed for the exploitation of the microviridin biosynthetic machinery for the production of novel variants through the establishment of an efficient one-pot in vitro platform. The relevance of this chemoenzymatic approach has been exemplified by the synthesis of novel potent serine protease inhibitors from both rationally-designed peptide libraries and bioinformatically predicted microviridins. Additionally, new structure-activity relationships (SARs) could be inferred by screening microviridin intermediates. The significance of this technique was further demonstrated by the simple incorporation of ncAAs into the microviridin scaffold.},
  language  = {en}
}
@article{MeyerMainzKehretal.2017,
  author    = {Meyer, Sabine and Mainz, Andi and Kehr, Jan-Christoph and Suessmuth, Roderich and Dittmann, Elke},
  title     = {Prerequisites of Isopeptide Bond Formation in Microcystin Biosynthesis},
  series = {ChemBioChem : a European journal of chemical biology},
  volume    = {18},
  journal   = {ChemBioChem : a European journal of chemical biology},
  publisher = {Wiley-VCH},
  address   = {Weinheim},
  issn      = {1439-4227},
  doi       = {10.1002/cbic.201700389},
  pages     = {2376 -- 2379},
  year      = {2017},
  abstract  = {The biosynthesis of the potent cyanobacterial hepatotoxin microcystin involves isopeptide bond formation through the carboxylic acid side chains of d-glutamate and -methyl d-aspartate. Analysis of the in vitro activation profiles of the two corresponding adenylation domains, McyE-A and McyB-A(2), either in a didomain or a tridomain context with the cognate thiolation domain and the upstream condensation domain revealed that substrate activation of both domains strictly depended on the presence of the condensation domains. We further identified two key amino acids in the binding pockets of both adenylation domains that could serve as a bioinformatic signature of isopeptide bond-forming modules incorporating d-glutamate or d-aspartate. Our findings further contribute to the understanding of the multifaceted role of condensation domains in nonribosomal peptide synthetase assembly lines.},
  language  = {en}
}