Applications of Mass Spectrometry to Membrane Protein Drug Development

Membrane proteins constitute the largest class of current drug targets. With many atomic structures now coming to the fore, including human protein targets, we have an unprecedented opportunity to study and rationalise the action of drugs on these important targets. One of the reasons why structures of these important proteins are only just becoming available is the inherent properties of these complexes that have hindered their study over the years; specifically their low expression levels and poor solubility contribute to the practical challenges that had to be overcome. Now that atomic structures and mechanistic insights are emerging,new methods are therefore required to assess their drug-binding properties and to contribute to data gleaned from other biophysical methods. Mass spectrometry is one such method that is providing fascinating insight into the properties of membrane proteins. Without the complication of protective coatings, required for solubility, the gas phase can be an ideal environment for the study membrane proteins, liberated from the bilayer, but often with critical lipid binding maintained. We are keen to develop further to our initial findings that lipids can modulate the properties of membrane proteins by extending these observations to the role of lipids and drugs simultaneously. We have selected three different themes, all of high strategic importance for drug development, described briefly here. The ability of bacteria to resist the challenge of antibiotics is a global threat to human health. Bacteria use a variety of mechanisms to do this including through pumps embedded in cell membranes to actively expel drugs or by controlling import of drugs through pores, formed by proteins in their outer cell-envelopes. One objective of our research programme is to apply mass spectrometry to gain new insight into these two mechanisms. Specifically, we will study the roles of lipids that play in holding open pores for drug import and in speeding up the transport of drugs out through pumps. Since many drugs now target membrane proteins a plethora of unwanted side effects are occurring through indiscriminate binding to other membrane proteins. These include binding of antipsychotic drugs to the human glucose transporter, anti HIV drugs to proteins involved in ageing and anticancer drugs to multidrug resistance pumps and to a human transporter of unknown function. A second objective is therefore to develop and apply mass spectrometry methods to study the unwanted side effects of drug binding to these targets. A third objective of our research programme involves the study of complexes involved in the assembly and synthesis of the armoury that protects the bacterial cell envelope. Among these complexes, one folds and assembles the pore proteins and the other orchestrates the formation of peptidoglycan layer. Both are therefore attractive targets for antibiotic intervention. For example, if we could uncover new targets that prevent folding and assembly of the pore proteins we could increase their synthesis to improve drug import. Similarly if we could control synthesis of the protective peptidoglycan layer we could affect the survival of bacteria, their division and the import of drugs. Overall therefore this research programme will have a number of beneficiaries. Firstly, it will benefit those using mass spectrometry by uncovering new methods to gain insight into the structural biology of membrane proteins. Secondly, the research outcomes will be of interest to pharmaceutical companies and biotechnology industries, particularly those with a focus on membrane protein as targets in human health or nanotechnology devices. In the longer-term interpreting mechanisms of drug resistance, unravelling the unwanted side-effects of drugs and understanding of the cell-wall biosynthesis of pathogenic bacteria will impact global human health.