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Structure and supramolecular organization of membrane transport proteins

Applicant Fotiadis Dimitrios
Number 144168
Funding scheme Project funding (Div. I-III)
Research institution Institut für Biochemie und Molekulare Medizin Universität Bern
Institution of higher education University of Berne - BE
Main discipline Biophysics
Start/End 01.11.2012 - 31.10.2015
Approved amount 430'000.00
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All Disciplines (2)

Discipline
Biophysics
Biochemistry

Keywords (13)

transmission electron microscopy; crystallization; atomic force microscopy; two-dimensional crystal; amino acid transport protein; membrane transport protein; X-ray crystallography; membrane protein; three-dimensional crystal; electron crystallography; glucose transport protein; structure; cation-chloride cotransporter

Lay Summary (English)

Lead
Lay summary

Transport proteins are the gatekeepers of cells and cell organelles, and are involved in cellular nutrition, ionic homeostasis, disposal of waste products and uptake of certain drugs. Dysfunction of these membrane proteins leads to a number of serious human diseases. Yet the structural information required to help us understand cellular transport and genetic disorders, and design effective drugs is lacking. High-resolution structures of pro- and eukaryotic transport proteins, and indeed of membrane proteins in general, are sparse compared to structures of water-soluble proteins. The reasons for this are mainly attributed to the amphiphilic nature of membrane proteins and the difficulty to overexpress them heterologously.

This Swiss National Science Foundation (SNSF) project proposal aims to elucidate the structure, the conformational changes upon substrate binding and the supramolecular organization of selected prokaryotic and mammalian transport proteins by X-ray crystallography, cryo-transmission electron microscopy and atomic force microscopy. Certain projects are continuations of our previous SNSF proposal (No. 31003A_125150), while others are new. Promising preliminary results are presented and form the basis of this new SNSF proposal.

Our target transport proteins are prokaryotic and human members of the amino acid/polyamine/organocation superfamily, including two members of the L-type amino acid transporter (LAT) subfamily, the glucose phosphoenolpyruvate: carbohydrate phosphotransferase system transporter from Escherichia coli, and the potassium-chloride cotransporter KCC4 from mouse. We address important open questions from the membrane transport protein field and aim to answer them by determining the low-, medium- and high-resolution structures of our target proteins. These questions include the structures and conformations of antiporters in their native environment, the lipid bilayer (in contrast to detergent-solubilized proteins), the structure and supramolecular organization of human heteromeric amino acid transporters (HATs), the molecular basis of glucose binding and transport, and the supramolecular structure of mammalian cation-chloride cotransporters.

Knowledge of these target structures will increase our general understanding of transporter organization and molecular transport mechanism. Furthermore, 3D structures at high-resolution, e.g. of human HAT and prokaryotic LAT, which has high amino acid identity to human LATs, will facilitate the development of new inhibitors for pharmaceutical applications by structure-based drug design.


Direct link to Lay Summary Last update: 21.02.2013

Responsible applicant and co-applicants

Employees

Publications

Publication
2D and 3D crystallization of the wild-type IIC domain of the glucose PTS transporter from Escherichia coli
Kalbermatter David, Jeckelmann Jean Marc, Chiu Po Lin, Ucurum Zöhre, Walz Thomas, Fotiadis Dimitrios (2014), 2D and 3D crystallization of the wild-type IIC domain of the glucose PTS transporter from Escherichia coli, in Journal of Structural Biology, 191(3), 376-380.
Cultivation strategies to enhance productivity of Pichia pastoris: A review
Looser V., Brühlmann B., Bumbak F., Stenger C., Costa M., Camattari A., Fotiadis D., Kovar K. (2014), Cultivation strategies to enhance productivity of Pichia pastoris: A review, in Biotechnology Advances, 1177.
Detergent-Induced Stabilization and Improved 3D Map of the Human Heteromeric Amino Acid Transporter 4F2hc-LAT2
Meury Marcel, Costa Meritxell, Harder Daniel, Stauffer Mirko, Jeckelmann Jean-Marc, Bruehlmann Bela, Rosell Albert, Ilgue Hueseyin, Kovar Karin, Palacin Manuel, Fotiadis Dimitrios (2014), Detergent-Induced Stabilization and Improved 3D Map of the Human Heteromeric Amino Acid Transporter 4F2hc-LAT2, in PLOS ONE, 9(10), e109882.
Structural bases for the interaction and stabilization of the human amino acid transporter LAT2 with its ancillary protein 4F2hc
Rosell Albert, Meury Marcel, Alvarez-Marimon Elena, Costa Meritxell, Perez-Cano Laura, Zorzano Antonio, Fernandez-Recio Juan, Palacin Manuel, Fotiadis Dimitrios (2014), Structural bases for the interaction and stabilization of the human amino acid transporter LAT2 with its ancillary protein 4F2hc, in PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA, 111(8), 2966-2971.

Collaboration

Group / person Country
Types of collaboration
Prof. Manuel Palacin / University of Barcelona Spain (Europe)
- in-depth/constructive exchanges on approaches, methods or results
- Publication
- Exchange of personnel

Scientific events

Active participation

Title Type of contribution Title of article or contribution Date Place Persons involved
Research Symposium in Honor of Prof. Krzysztof Palczewski Talk given at a conference From receptors to transporters 19.10.2015 Cleveland, OH, United States of America Fotiadis Dimitrios; Harder Daniel;
Seminar at TUM Individual talk Role of electrostatic interactions for ligand recognition and specificity of peptide transporters 03.03.2015 Munich, Germany Fotiadis Dimitrios; Harder Daniel;
Scientific opportunities in protein 2D-crystallography at (Swiss)FELs Talk given at a conference Atomic Force Microscopy for the Study of Membrane Proteins 27.01.2015 Paul Scherrer Institute, Villigen, Switzerland Kalbermatter David; Fotiadis Dimitrios;
BioTech 2014 Talk given at a conference Structure and Function of a Human Heteromeric Amino Acid Transporter 13.06.2014 Prague, Czech Republic Costa Torres Meritxell; Fotiadis Dimitrios; Harder Daniel;
Seminar at ZHAW Individual talk Structural Biology of Membrane Proteins 21.12.2012 Wädenswil, Switzerland Costa Torres Meritxell; Fotiadis Dimitrios;


Communication with the public

Communication Title Media Place Year
Media relations: print media, online media Crystal clear International Innovation International 2013
Media relations: print media, online media Membrane proteins - at the core of life Public Service Review: European Science and Technology International 2012

Associated projects

Number Title Start Funding scheme
157704 Direct electron detector and phase plate for cryo-transmission electron microscopy of biological samples 01.12.2014 R'EQUIP
125762 NCCR TransCure: From transport physiology to identification of therapeutic targets (phase I) 01.11.2010 National Centres of Competence in Research (NCCRs)
162581 Structure and supramolecular organization of membrane transport proteins 01.12.2015 Project funding (Div. I-III)
125150 Structure and supramolecular organization of amino acid and peptide transport proteins 01.05.2009 Project funding (Div. I-III)
125150 Structure and supramolecular organization of amino acid and peptide transport proteins 01.05.2009 Project funding (Div. I-III)

Abstract

Transport proteins are the gatekeepers of cells and cell organelles, and are involved in cellular nutrition, ionic homeostasis, disposal of waste products and uptake of certain drugs. Dysfunction of these membrane proteins leads to a number of serious human diseases. Yet the structural information required to help us understand cellular transport and genetic disorders, and design effective drugs is lacking. High-resolution structures of pro- and eukaryotic transport proteins, and indeed of membrane proteins in general, are sparse compared to structures of water-soluble proteins. The reasons for this are mainly attributed to the amphiphilic nature of membrane proteins and the difficulty to overexpress them heterologously.This Swiss National Science Foundation (SNSF) project proposal aims to elucidate the structure, the conformational changes upon substrate binding and the supramolecular organization of selected prokaryotic and mammalian transport proteins by X-ray crystallography, cryo-transmission electron microscopy and atomic force microscopy. Certain projects are continuations of our previous SNSF proposal (No. 31003A_125150), while others are new. Promising preliminary results are presented and form the basis of this new SNSF proposal.Our target transport proteins are prokaryotic and human members of the amino acid/polyamine/organocation superfamily, including two members of the L-type amino acid transporter (LAT) subfamily, the glucose phosphoenolpyruvate: carbohydrate phosphotransferase system transporter from Escherichia coli, and the potassium-chloride cotransporter KCC4 from mouse. We address important open questions from the membrane transport protein field and aim to answer them by determining the low-, medium- and high-resolution structures of our target proteins. These questions include the structures and conformations of antiporters in their native environment, the lipid bilayer (in contrast to detergent-solubilized proteins), the structure and supramolecular organization of human heteromeric amino acid transporters (HATs), the molecular basis of glucose binding and transport, and the supramolecular structure of mammalian cation-chloride cotransporters.Knowledge of these target structures will increase our general understanding of transporter organization and molecular transport mechanism. Furthermore, 3D structures at high-resolution, e.g. of human HAT and prokaryotic LAT, which has high amino acid identity to human LATs, will facilitate the development of new inhibitors for pharmaceutical applications by structure-based drug design.
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