Engineering of Biomolecular Energy Conversion and Transport Modules for the Assembly of Molecular Factories

Schematic representation of a ‘simple’ biomolecular factory: The reactor is a nanocontainer (made of lipid or block copolymer) equipped with light-driven proton pumps (in red; energizing modules), proton-driven solute transporters (in blue; translocating modules) and metabolizing enzymes (in brown; metabolizing modules). Such light powered nanoreactors will import specific solutes from a solution and degrade them inside. Possible application: decontamination of water/solutions from toxic compounds or pollutant.

Building autonomous synthetic organelles and cells with a defined function using a repertoire of functional modules (toolkit) and containing inside a minimal metabolism for survival, represents the ultimate goal of this project group.

Such complex processors will open a wide variety of possibilities ranging from environmental to medical applications. One of the most important challenges will be to provide a large repertoire of engineered and modular biomolecular-transport and -energy conversion systems for assembly of nanoreactors with diverse functionalities in lipid bilayers and block copolymers.

Initially, modules will include light-driven proton pumps and proton-driven solute transporters in the membrane, and metabolizing enzymes inside the container. Next, more complex powering systems will be explored such as combinations of light-driven proton pumps with sodium/proton antiporters with the objective: to energise sodium-driven solute transporters. This will significantly increase the repertoire and specificity of translocating modules.

The availability of numerous, highly specialized membrane proteins in milligram amounts offers the unique opportunity to use them as building blocks and toolkit to assemble molecular factories in the form of nanoreactors and functional surfaces using bottom-up approaches.

This project group has a strong expertise and knowledge in biochemistry, function and structure of membrane proteins. Furthermore, the group already possesses a significant number of recombinant transport proteins for different solutes such as peptides, sugars, amino acids and antibiotics that can be used as modules for engineering and assembly of nanoreactors.

Publications

N. Ayoub, P. Roth, Z. Ucurum, D. Fotiadis, S. Hirschi “Structural and biochemical insights into His-tag-induced higher-order oligomerization of membrane proteins by cryo-EM and size exclusion chromatography“ J. Struct. Biol. 2023. [DOI]

S. Hirschi, T. R. Ward, W. P. Meier, D. J. Müller, D. Fotiadis “Synthetic Biology: Bottom-Up Assembly of Molecular Systems“ Chem. Rev. 2022. [DOI]

M. Stauffer, J. Jeckelmann, H. Ilgü, Z. Ucurum, R. Boggavarapu, D. Fotiadis “Peptide transporter structure reveals binding and action mechanism of a potent PEPT1 and PEPT2 inhibitor“ Commun. Chem. 2022. [DOI]

F. Ruggeri, C. Schwemmer, M. Stauffer, P. M. Nicollier, J. Figueiredo da Silva, P. D. Bosshart, K. Kochems, D. Fotiadis, A. Knoll, H. Wolf “Placement of Biological Membrane Patches in a Nanofluidic Gap With Control Over Position and Orientation“ 2022. [DOI]

M. Stauffer, Z. Ucurum, D. Harder, D. Fotiadis “Engineering and functional characterization of a proton-driven β-lactam antibiotic translocation module for bionanotechnological applications“ Sci. Rep. 2021. [DOI]

S. Hirschi, D. Kalbermatter, Z. Ucurum, T. Lemmin, D. Fotiadis “Cryo-EM structure and dynamics of the green-light absorbing proteorhodopsin“ Nat. Commun. 2021. [DOI]

M. Stauffer, S. Hirschi, Z. Ucurum, D. Harder, R. Schlesinger, D. Fotiadis “Engineering and Production of the Light-Driven Proton Pump Bacteriorhodopsin in 2D Crystals for Basic Research and Applied Technologies“ Methods Protoc. 2020. [DOI]

S. Hirschi, D. Kalbermatter, Z. Ucurum, D. Fotiadis “Cryo-electron microscopic and X-ray crystallographic analysis of the light-driven proton pump proteorhodopsin reveals a pentameric assembly“ J. Struct. Biol. X 2020. [DOI]

S. Hirschi, D. Fotiadis “Purification of Membrane Proteins by Affinity Chromatography with On-Column Protease Cleavage“ Expression, Purification, and Structural Biology of Membrane Proteins 2020. [DOI]

J. Gaitzsch, S. Hirschi, S. Freimann, D. Fotiadis, W. Meier “Directed Insertion of Light-Activated Proteorhodopsin into Asymmetric Polymersomes from an ABC Block Copolymer“ Nano Lett. 2019. [DOI]

S. Hirschi, N. Fischer, D. Kalbermatter, P. R. Laskowski, Z. Ucurum, D. J. Müller, D. Fotiadis “Design and assembly of a chemically switchable and fluorescently traceable light-driven proton pump system for bionanotechnological applications“ Sci. Rep. 2019. [DOI]

D. Kalbermatter, N. Shrestha, N. Ader-Ebert, M. Herren, P. Moll, R. K. Plemper, K. Altmann, J. P. Langedijk, F. Gall, U. Lindenmann, R. Riedl, D. Fotiadis, P. Plattet “Primary Resistance Mechanism of the Canine Distemper Virus Fusion Protein against a Small-Molecule Membrane Fusion Inhibitor“ Virus Res. 2018. [DOI]

R. Goers, J. Thoma, N. Ritzmann, A. Di Silvestro, C. Alter, G. Gunkel-Grabole, D. Fotiadis, D. J. Müller, W. Meier “Optimized reconstitution of membrane proteins into synthetic membranes“ Commun. Chem. 2018. [DOI]

J. Thoma, S. Manioglu, D. Kalbermatter, P. D. Bosshart, D. Fotiadis, D. J. Müller “Protein-enriched outer membrane vesicles as a native platform for outer membrane protein studies“ Commun. Biol. 2018. [DOI]

P. R. Laskowski, M. Pfreundschuh, M. Stauffer, Z. Ucurum, D. Fotiadis, D. J. Müller “High-Resolution Imaging and Multiparametric Characterization of Native Membranes by Combining Confocal Microscopy and Atomic Force Microscopy-Based Multifunctional Toolbox“ ACS Nano 2017. [DOI]

W. Aeschimann, S. Staats, S. Kammer, N. Olieric, J. Jeckelmann, D. Fotiadis, T. Netscher, G. Rimbach, M. Cascella, A. Stocker “Self-assembled α-Tocopherol Transfer Protein Nanoparticles Promote Vitamin E Delivery Across an Endothelial Barrier“ Sci. Rep. 2017. [DOI]

N. Ritzmann, J. Thoma, S. Hirschi, D. Kalbermatter, D. Fotiadis, D. J. Müller “Fusion Domains Guide the Oriented Insertion of Light-Driven Proton Pumps into Liposomes“ Biophys J. 2017. [DOI]

D. Kalbermatter, P. Chiu, J. Jeckelmann, Z. Ucurum, T. Walz, D. Fotiadis “Electron crystallography reveals that substrate release from the PTS IIC glucose transporter is coupled to a subtle conformational change“ J. Struct. Biol. 2017. [DOI]

P. Orekhov, A. Bothe, H. Steinhoff, K. V. Shaitan, S. Raunser, D. Fotiadis, R. Schlesinger, J. P. Klare, M. Engelhard “Sensory Rhodopsin I and Sensory Rhodopsin II Form Trimers of Dimers in Complex with their Cognate Transducers“ Photochem. Photobiol. 2017. [DOI]

M. Pfreundschuh, D. Harder, Z. Ucurum, D. Fotiadis, D. J. Müller “Detecting Ligand-Binding Events and Free Energy Landscape while Imaging Membrane Receptors at Subnanometer Resolution“ Nano Lett. 2017, 17(5):3261-69. [DOI]

D. Harder, S. Hirschi, Z. Ucurum, R. Goers, W. Meier, D. J. Müller, D. Fotiadis “Engineering a Chemical Switch into the Light-driven Proton Pump Proteorhodopsin by Cysteine Mutagenesis and Thiol Modification“ Angew. Chem. Int. Ed. 2016, 55:8846. [DOI]

S. Hirschi, M. Stauffer, D. Harder, D. J. Müller, W. Meier, D. Fotiadis “Engineering and Assembly of Protein Modules into Functional Molecular Systems“ Chimia 2016, 6:398. [DOI]

R. Boggavarapu, S. Hirschi, D. Harder, M. Meury, Z. Ucurum, M. J. Bergeron, D. Fotiadis “Purification of Human and Mammalian Membrane Proteins Expressed in Xenopus laevis Frog Oocytes for Structural Studies“ Heterologous Expression of Membrane Proteins 2016, 1432:223-42. [DOI]

D. Kalbermatter, J. Jeckelmann, P. Chiu, Z. Ucurum, T. Walz, D. Fotiadis “2D and 3D crystallization of the wild-type IIC domain of the glucose PTS transporter from Escherichia coli“ J. Struct. Biol. 2015, 191:376-80. [DOI]

R. Boggavarapu, J. Jeckelmann, D. Harder, Z. Ucurum, D. Fotiadis “Role of electrostatic interactions for ligand recognition and specificity of peptide transporters“ BMC Biol. 2015, 13:DOI: 10.1186/s12915-015-0167-8. [DOI] [More Information]

D. Fotiadis, A. Engel “Two‐Dimensional Crystallisation of Membrane Proteins and Structural Assessment“ eLS 2015:1-10. [DOI]

R. Petrosyan, C. A. Bippes, S. Walheim, D. Harder, D. Fotiadis, T. Schimmel, D. Alsteens, D. J. Müller “Single-Molecule Force Spectroscopy of Membrane Proteins from Membranes Freely Spanning Across Nanoscopic Pores“ Nano Lett. 2015, 15:3624. [DOI]

P. D. Bosshart, A. Engel, D. Fotiadis “High-Resolution Atomic Force Microscopy Imaging of Rhodopsin in Rod Outer Segment Disk Membranes“ Rhodopsin 2015:189-203. [DOI]

Project Leader

Dimitrios Fotiadis

Persons involved

Stephan Hirschi
Mirko Stauffer

Lab

Fotiadis group @UniBe