Technical University of Berlin View Institution's Website 6 articles published in JoVE Bioengineering Antimicrobial Peptides Produced by Selective Pressure Incorporation of Non-canonical Amino Acids Jessica H. Nickling*1, Tobias Baumann*1, Franz-Josef Schmitt2, Maike Bartholomae3, Oscar P. Kuipers3, Thomas Friedrich2, Nediljko Budisa1 1Department of Biocatalysis, Institute of Chemistry, Technische Universität Berlin, 2Department of Bioenergetics, Institute of Chemistry, Technische Universität Berlin, 3Molecular Genetics Group, Groningen Biomolecular Sciences and Biotechnology Institute, Department of Molecular Genetics, University of Groningen The protocol presents the Escherichia coli-based selective pressure incorporation of non-canonical amino acids (ncAAs) into the lactococcal antimicrobial peptide nisin. Its properties can be changed during recombinant expression via substitution with desired ncAAs in defined growth media. Resulting changes in bioactivity are mapped by growth inhibition assays and fluorescence microscopy. Bioengineering Engineering 'Golden' Fluorescence by Selective Pressure Incorporation of Non-canonical Amino Acids and Protein Analysis by Mass Spectrometry and Fluorescence Tobias Baumann1, Franz-Josef Schmitt2, Almut Pelzer1, Vivian Jeanette Spiering3, Georg Johannes Freiherr von Sass1, Thomas Friedrich2, Nediljko Budisa1 1Institute of Chemistry L 1, Department of Biocatalysis, Technical University of Berlin, 2Institute of Chemistry PC 14, Department of Bioenergetics, Technical University of Berlin, 3Institute of Chemistry TC 7, Department of Physical Chemistry/Molecular Material Sciences, Technical University of Berlin Synthetic biology enables the engineering of proteins with unprecedented properties using the co-translational insertion of non-canonical amino acids. Here, we presented how a spectrally red-shifted variant of a GFP-type fluorophore with novel fluorescence spectroscopic properties, termed "gold" fluorescent protein (GdFP), is produced in E. coli via selective pressure incorporation (SPI). Bioengineering A Method for Determination and Simulation of Permeability and Diffusion in a 3D Tissue Model in a Membrane Insert System for Multi-well Plates Hao-Hsiang Hsu1, John-Kevin Kracht1, Laura Elisabeth Harder1, Kerstin Rudnik1, Gerd Lindner2, Katharina Schimek2, Uwe Marx3, Ralf Pörtner1 1Institute of Bioprocess and Biosystems Engineering, Hamburg University of Technology, 2Institute of Biotechnology, Department Medical Biotechnology, Technische Universität Berlin, 3TissUse GmbH A method for determination of permeability in a membrane insert system for multi-well plates and in silico parameter optimization for the calculation of diffusion coefficients using simulation are presented. Cancer Research Study of Viral Vectors in a Three-dimensional Liver Model Repopulated with the Human Hepatocellular Carcinoma Cell Line HepG2 Thomas Hiller1, Viola Röhrs1, Eva-Maria Dehne2, Anke Wagner1,3, Henry Fechner1, Roland Lauster2, Jens Kurreck1 1Department of Applied Biochemistry, Institute of Biotechnology, Berlin University of Technology, 2Department of Medical Biotechnology, Institute of Biotechnology, Berlin University of Technology, 3Department of Bioprocess Engineering, Institute of Biotechnology, Berlin University of Technology The recellularized extracellular matrix of a decellularized rat liver can be used as a humanized, three-dimensional ex vivo model to study the distribution and transgene expression of a virus or viral vector. Bioengineering The Multi-organ Chip - A Microfluidic Platform for Long-term Multi-tissue Coculture Eva-Maria Materne*1, Ilka Maschmeyer*1,2, Alexandra K. Lorenz1, Reyk Horland1,2, Katharina M. S. Schimek1, Mathias Busek3, Frank Sonntag3, Roland Lauster1, Uwe Marx1,2 1Medical Biotechnology, Technische Universität Berlin, 2TissUse GmbH, 3Fraunhofer IWS Here, we present a protocol to coculture primary cells, tissue models and punch biopsies in a microfluidic multi-organ chip for up to 28 days. Human dermal microvascular endothelial cells, liver aggregates and skin biopsies were successfully combined in a common media circulation. Biology Measuring Cation Transport by Na,K- and H,K-ATPase in Xenopus Oocytes by Atomic Absorption Spectrophotometry: An Alternative to Radioisotope Assays Katharina L. Dürr1,2, Neslihan N. Tavraz1, Susan Spiller1, Thomas Friedrich1 1Institute of Chemistry, Technical University of Berlin, 2The Vollum Institute, Oregon Health & Science University We describe a method to quantify the activity of K+-countertransporting P-type ATPases by heterologous expression of the enzymes in Xenopus oocytes and measuring Rb+ or Li+ uptake into individual cells by atomic absorption spectrophotometry. The method is a sensitive and safe alternative to radioisotope flux experiments facilitating complex kinetic studies.