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Switchable transport across genetically engineered channel proteins

English title Switchable transport across genetically engineered channel proteins
Applicant Fischer Ozana
Number 132240
Funding scheme Ambizione
Research institution Physikalische Chemie Departement Chemie Universität Basel
Institution of higher education University of Basel - BS
Main discipline Physical Chemistry
Start/End 01.03.2011 - 28.02.2014
Approved amount 391'292.00
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Keywords (4)

polymer nanovesicles; switchable transport; engineered channel proteins; block copolymers

Lay Summary (English)

Lead
Lay summary
The aim of this project is to design and engineer channel proteins that can act as controllable gates once embedded in the membrane of nanovesicles. The reversible control of the transport of molecules through the channel protein can be established by changes in pH, temperature or light.

OmpF channel protein has been chosen as model channel protein due to its thermal and chemical stability. The OmpF channel protein will be equipped with polymers that are pH or temperature sensitive. The reversibility of the switch introduced inside the channel proteins' pores will be characterized once the channel proteins are embedded in the wall of polymer nanovesicles. The polymeric nanovesicles are created by self-assembly of amphiphilic block copolymers in water. Amphiphilic block copolymers are made of a hydrophobic block and a hydrophilic block and they spontaneously form vesicles in water. The polymeric systems are a novel alternative to the liposome carriers, and are more stable, while preserving all the advantages of lipidic systems, such as lack of immunogenicity. The obtained vesicles reconstituting engineered channel proteins will be characterized using fluorescence and spectroscopic methods. The pH/temperature-sensitive OmpF hybrid system will be a first step in mimicking nature (e.g. ligand gated channels like acetylcholine receptors, light gated like channel rhodopsin) to produce functional proteins with potential applications in biotechnology or biomedicine.

We anticipate that channel proteins engineered for reversible transport induced by changes in external stimuli (pH, temperature, light) will be used for tunable release of the content of liposomes or polymeric vesicles. This general system will be of special interest for various biomedical, biotechnological and biosensing applications. A biomedical application would be polymeric vesicles containing drugs or drug producing enzymes inside and embedded channel proteins that allow switchable transport of the prodrug/drug across the polymer membrane. With this specific system a precise control of the rate and the moment of the release of the drugs could be achieved.
Direct link to Lay Summary Last update: 21.02.2013

Responsible applicant and co-applicants

Employees

Publications

Publication
Reconstitution of OmpF membrane protein on bended lipid bilayers: perforated hexagonal mesophases.
Alexandru Zabara, Renatta Negrini, Patric Bauman, Ozana Fischer, Raffaele Mezzenga (2014), Reconstitution of OmpF membrane protein on bended lipid bilayers: perforated hexagonal mesophases., in Chemical Communication, 50(20), 2642-2645.
Light-responsive polymer nanoreactors: a source of reactive oxygen species on demand
Baumman Patric, Fischer-Onaca Ozana, Balasubramanian Vimalkumar, Sienkiewicz Andrej, Palivan Cornelia (2013), Light-responsive polymer nanoreactors: a source of reactive oxygen species on demand, in Nanoscale, 5, 217-224.
Polymeric Nanocarriers and Nanoreactors: A Survey of Possible Therapeutic Applications
Onaca-Fischer Ozana, Lui Juan, Inglin Mark, Palivan Cornelia (2012), Polymeric Nanocarriers and Nanoreactors: A Survey of Possible Therapeutic Applications, in Current Pharmaceutical Design, 18(18), 2622-2643.
Can polymeric vesicles that confine enzymatic reactions act as simplified organelles?
Pascal Tanner, Stefan Egli, Vimalkumar Balasubramanian, Ozana Onaca, Cornelia Palivan, Wolfgang Meier (2011), Can polymeric vesicles that confine enzymatic reactions act as simplified organelles?, in FEBS Letters , 585(11), 1699-1706.
Polymeric vesicles: From Drug Carriers to Nanoreactors and Artificial Organelles
Pascal Tanner, Patric Baumann, Ramona Enea, Ozana Onaca-Fischer, Cornelia Palivan, Wolfgang Meier (2011), Polymeric vesicles: From Drug Carriers to Nanoreactors and Artificial Organelles, in Accounts of Chemical Research , 44(10), 1039-1049.
Protein–polymer nanoreactors for medical applications
Cornelia Palivan, Ozana Fischer-Onaca, Mihaela Delcea, Fabian Itel, Wolfgang Meier (2011), Protein–polymer nanoreactors for medical applications, in Chemical Society Reviews, 41, 2800-2023.
Selective and Responsive Nanoreactors
Kasper Renggli, Patric Baumann, Karolina Langowska, Ozana Onaca, Nico Bruns, Wolfgang Meier (2011), Selective and Responsive Nanoreactors, in Adv. Funct. Mater, 21(7), 1241-1259.
Intercompartmental enzymatic cascade in channel-equipped polymersome-in-polymersome architectures
Winna Siti, Hans-Peter Michael De Hoog, Ozana Onaca Fischer, Nikodem Tomczak, Yee Shan Wong, Madhavan Nallani, Bo Liedberg, Intercompartmental enzymatic cascade in channel-equipped polymersome-in-polymersome architectures, in J. Mater. Chem. B.
Nanoreactors for Biomedical Applications
Itel Fabian, Dinu Adrian, Tanner Pascal, Fischer Ozana, Palivan Cornelia, Nanoreactors for Biomedical Applications, in Torchilin Vladimir (ed.), World Scientific Publishing Co, Announced after publication.
Perforated Bicontinuous Cubic Phases with pH-Responsive Topological Channel Interconnectivity
Zabara Alexandru, Fischer-Onaca Ozana, Mezzenga Raffaele, Perforated Bicontinuous Cubic Phases with pH-Responsive Topological Channel Interconnectivity, in Small.
Polymer Nanoreactors
Edlinger Christoph, Yang Xiaoyan, Fischer-Onaca Ozana, Palivan Cornelia, Polymer Nanoreactors, in Mihai Peterca (ed.), John Wiley and Sons, Inc, New Jersey.

Collaboration

Group / person Country
Types of collaboration
Nanyang Technological University Singapore (Asia)
- Publication
ETH Switzerland (Europe)
- in-depth/constructive exchanges on approaches, methods or results
- Publication

Scientific events

Active participation

Title Type of contribution Title of article or contribution Date Place Persons involved
23th Liposome workshop Talk given at a conference Stimulus-responsive polymer nanoreactors for efficient photodynamic therapy 24.04.2014 Oberjoch, Germany Fischer Ozana;
247th ACS National Meeting & Exposition Talk given at a conference Stimulus-responsive polymer nanoreactors for efficient photodynamic therapy 16.03.2014 Dallas, United States of America Fischer Ozana;
ISN2A Poster Switchable Transport across Modified Membrane Channel Proteins 21.01.2014 Lisbon, Portugal Fischer Ozana;
ITRG,2012 Individual talk Controlling Memebrane Permeability with Channel Proteins 29.10.2012 Strasbourg, France Fischer Ozana;
Swiss Soft Days V Poster Controlling membrane permeability with channel proteins 08.06.2011 Basel, Switzerland Fischer Ozana;


Abstract

In biological membranes channel proteins facilitate the transport of different molecules into and out of the cell. The transport of substances can be active transport or passive diffusion. Channel proteins like OmpF and Opdk which are found in the outer membrane of Escherichia coli and Pseudomonas aeruginosa, respectively, act as passive gateways for different small molecules (less than 600 Da). However in biological cell membranes they do not undergo conformational changes for increased selectivity in molecule transport.The aim of the project is to engineer channel proteins in order to obtain a reversible switchable transport across them. To achieve this, we plan to modify such unspecific channel proteins by introducing cysteines into the genes of the channels proteins. Responsive polymer chains that can respond to external stimuli such as temperature, light, pH, presence of ions, will be chemically attached to or grown from the introduced cysteines in order to constrict the channel. Upon appropriate exterior input, the polymer chain will change its solubility resulting in blockage of the channel pore, while restoring the initial conditions will cause the polymer to assume its earlier form of solubility and thus reopening the channel protein. By modulation of the polymer chain length it is possible to control the cutoff of the pore, transforming it in a molecular sieve for substances in certain molecular weight ranges. To the best of our knowledge, the approach for triggered transport across channel proteins would be the first of its kind for biological pores and will therefore offer a general strategy for obtaining reversible biological pores. Moreover we envisage that a blockage of the channel proteins with sensitive peptides (pH, temperature) introduced using genetic engineering methods will offer an alternative to the use of synthetic polymers for blocking.The reversibility of the switch introduced inside the channel proteins` pores will be characterized once the channel proteins are reconstituted in polymer nanovesicles. The polymeric nanovesicles are created by self assembly of amphiphilic block copolymers, for e.g poly-(2-methyloxazoline)-poly (dimethylsiloxane)-poly (2-methyloxazoline). The polymeric systems are a novel alternative to the liposome carriers, and are more stable, while preserving all the advantages of lipidic systems, such as lack of immunogenicity. The polymeric vesicles reconstituting engineered channel proteins will be characterized using fluorescence and spectroscopic methods. We anticipate that channel proteins engineered for reversible, controlled transport caused by changes in external stimulation (pH, temperature, light) will be used in drug delivery for tunable release of the content of liposomes or polymeric vesicles, as well as in biotechnological applications or biosensing.
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