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Bionanoparticle formation for drug discovery

Primary Supervisor: Professor T.R. Dafforn, School of Biosciences

Secondary supervisor: Dr Sarah L. Horswell

PhD project title: Bionanoparticle formation for drug discovery

University of Registration: University of Birmingham

Project outline:

Biochemical and structural studies of membrane proteins has remained frustratingly challenging. For example, only 1 in every 50 protein structures are membrane proteins. This is in a background where understanding membrane protein function is fundamental to our understanding of biology, a fact that is mirrored by the observation that more than 50% of therapeutics and 70% of agrochemicals target membrane proteins.

One of the most important issues in studying membrane proteins is the process of extracting them from membranes in a form that is pure, active and stable.

In 2009 we developed a solution to this issue that allowed the extraction of membrane proteins from cell membranes complete with the local lipid environment. The method uses a polymer based on Styrene Maleic Acid (SMA) that excises a 10 nm diameter disc (SMALP) from cell membranes to produce a particle that contains a membrane protein in its local lipid environment.

We have shown that the method extracts a range of stabilised membrane proteins from a range of membranes. The method is now being adopted by academic and commercial scientists alike and has recently been used to produce both X-ray and cryo-EM structures of membrane proteins.

However, our understanding of the processes that underlie the formation of these SMALP particles is still limited. This gap in our knowledge becomes particularly important during the development of new polymers with improved activities and added functionalities. These polymers will not only provide better reagents for current uses of SMALPs but will also provide the foundation for an entirely new class of bionanoparticle that could be used in areas as wide ranging as sensing and energy production.

In this project we aim to carry out studies on the SMALP formation process using a range of biophysical techniques. In the first stage, we will:

  • Measure the kinetics of the process of SMALP formation from liposomes (using NMR and X-ray scattering) and from supported and floating bilayers (using capacitance and impedance measurements)
  • Measure kinetics of removal of lipids from floating monolayers (using Langmuir techniques)
  • Study the nature of the interaction between the polymers and the lipids (using vibrational spectroscopy to determine which functional groups are involved and how bilayer structure is perturbed, AFM to study the structural changes as the polymer interacts with a bilayer in the lateral direction and reflectivity to study these changes in the direction normal to the bilayer)

We will study these processes with a range of compositions typical of mammalian, bacterial, plant and fungal cell membranes to ascertain whether there are specific interactions that can be exploited for targeted use of the polymers.

In the second stage, we will employ similar strategies to study the interaction of the polymer with membrane proteins and smaller peptides. The purpose of these studies will be to find a means of predicting whether a protein is likely to retain its function when incorporated in a SMALP. In this stage we will also make use of circular dichroism and solution-based vibrational spectroscopy studies to measure changes in protein conformation.

Taken together these studies will provide insights that will help expand the use of SMALP technology in a wide range of commercial and academic applications.

BBSRC Strategic Research Priority: Understanding the Rules of Life: Structural Biology & Integrated Understanding of Health: Pharmaceuticals

Techniques that will be undertaken during the project:

  • Protein production through microbial expression
  • Membrane protein purification
  • Biophysical measurement including:
    • Circular Dichroism spectroscopy
    • Fluorescence spectroscopy
    • Fluorescence microscopy
    • Surface pressure-area isotherm measurements and monolayer transfer
    • Capacitance and impedance measurements
    • Interfacial vibrational spectroscopy (infrared and Raman)
    • Atomic force microscopy
    • NMR
    • Light scattering
    • Neutron and X-ray Scattering
    • Neutron and X-ray Reflectometry
  • Polymer synthesis and modification

Contact: Professor Tim Dafforn, University of Birmingham