Membrane proteins control the exchange of substances, the transfer of signals in the cells’ interior or external stimuli. What role do membrane proteins play in the pharmaceutical industry, for example as drug targets? Which problems need to be solved by biochemists in order to get these molecules under control? Dr. Uwe Schulte of the Freiburg-based biotech company Logopharm GmbH has amassed a great deal of experience in this field over the last few years, both in projects focusing on basic research as well as in projects carried out in cooperation with the pharmaceutical industry. Logopharm is a specialist in the analysis of membrane proteins, membrane protein complexes and functional networks involving membrane proteins. In an interview with BIOPRO, Schulte expresses his views on the direction research should take.
BIOPRO: Dr. Schulte, how important are membrane proteins for the pharmaceutical and biotechnology sectors?
Schulte: Until a couple of years ago, around two thirds of all drugs targeted membrane proteins, including receptors that transfer external signals into cells or ion channels that control the flow of current to nerve cells. Substances that target such membrane proteins have the potential to alter the behaviour of cancer cells or modulate the transfer of information between neurons. Since then, there has been major progress in the technologies that overcome membrane barriers, in the sense that a growing number of pharmaceutically active substances is now able to directly influence intracellular proteins, including enzymes or modulators of signalling networks. This is hugely advantageous because the characterisation of membrane proteins is far more difficult than the characterisation of proteins found in the aqueous environment inside cells.
BIOPRO: Does this mean that membrane proteins are no longer interesting targets for the pharmaceutical industry?
Schulte: No, it doesn’t. Membrane proteins still have huge potential. The majority of signalling pathways in a cell start at the receptor proteins in the membrane. In addition, specific membrane proteins on the surface of a certain cell type can be used as recognition structures. For example, this enables cancer drugs that are also toxic for healthy cells to be specifically directed to the malignant cells. Such surface markers can be used to visualise certain cell types or cellular modifications, something that is extremely important in the diagnosis of diseases. Membrane proteins and membrane protein complexes are definitely an area on which most research will concentrate in the next few years.
BIOPRO: Why do many pharmaceutical companies nevertheless not focus on membrane proteins?
Schulte: Membrane proteins are embedded in the cell membrane whose lipids create a specific hydrophobic environment. It is difficult to isolate these proteins and use standard biochemical methods to investigate them without interfering with their structure or composition. Moreover, many membrane proteins need to bind to specific lipids in the membrane to exert their physiological activity. One such example is certain types of potassium channels that bind to different forms of phosphatidylinositol that is essential for the transmission of signals. We are gradually shaking off the traditional view according to which the membrane is a pure solvent for proteins. Biological membranes are complex functional units and membrane proteins are only one component of biological membranes. They must not be looked at in isolation; complicated experimental set-ups are necessary. And another major, i.e. third aspect, relates to the concentration of plasma membrane proteins, which is lower than that of many other proteins. When researchers investigate them, they either need to be enriched or highly sensitive measurement methods need to be used.
BIOPRO: Logopharm GmbH have developed the so-called microproteomics method for this, haven't they?
Schulte: Yes, they have. It’s a technology platform that combines two components. One aspect of this platform is the optimisation of each individual step from the isolation of the proteins from biological membranes to their purification in order to achieve a quantity of protein that is as high as possible. We have also developed miniaturised devices and tools that are suitable for the smallest volume samples. Another aspect is the use of extremely precise and sensitive mass spectrometers, which enable us to detect and quantitatively determine the smallest amounts of proteins in a sample. These methods are suitable for investigating membrane proteins from very small tissue quantities or rare cell types.
BIOPRO: Can you give me an example of a project that you have carried out in cooperation with or on behalf of the pharmaceutical industry?
Schulte: For example, we have examined the molecular composition of the target structure of the cholesterol-lowering drug Ezetimide on behalf of Merck. At the time we did the investigation, the drug was already on the market, but no details about the target protein were available and neither was it known whether other proteins contributed to its effect. We identified a membrane transporter protein as the direct binding partner and we were able to exclude the involvement of other proteins in this process. This was an important step in clarifying the mechanism of the drug’s action and is the basis for the specific development of new drugs that target this membrane transporter.
BIOPRO: In which direction is research on membrane proteins going?
Schulte: We now know that membrane proteins do not exert their function as individual proteins. They are frequently associated with other membrane proteins or with proteins in the cell's interior in the form of complexes or networks. If one seeks to understand and modulate their function, it is necessary to decipher these complexes and networks. In cooperation with the research group led by Prof. Dr. Bernd Fakler at the Institute of Physiology at the University of Freiburg, we have for example investigated a family of calcium channels in neurons. These channels interact with a broad range of proteins, which in turn are associated with other proteins; we have identified around two hundred proteins in this network. We also used our methods to determine the quantity of the individual proteins in order to find out which interactions are particularly stable or frequent, and in order to identify protein complexes in these networks. In principle, it is difficult to automate such interaction studies with biological tissue, therefore we do not offer high-throughput experiments. Our customers benefit from our support in solving very complicated issues, where we need to look closely into all the aspects and which require the highest technical quality.
BIOPRO: How has Logopharm developed over the last few years?
Schulte: Overall, Logopharm has developed in a fairly positive way; we have been in the black since last year and our customers come from industry and academia all over the world. And as I said previously: it will become increasingly important to develop drugs that do not target individual proteins. The pharmaceutical industry will need to increasingly focus on the interactions of proteins in complexes and networks in order to be able to develop more effective and selective therapies. This requires specialised and research-oriented service providers such as Logopharm GmbH.
Dr. Uwe Schulte