Part II Undergraduate Student, University of Oxford, 2001–2002
D.Phil. Student, University of Oxford, 2002–present
Spectroscopy of Supported Single Lipid Bilayers and Peptides
I use surface-sensitive techniques to study the interaction of membrane-inserting peptides with fully hydrated solid-supported single phospholipid bilayers. The main peptide I have studied is indolicidin, a naturally occurring antimicrobial peptide. Its cytotoxicity at low concentrations is partly from disruption to the cell membrane and partly due to attack of the nucleus after stealthy membrane penetration. To address its effect on the lipids, I study its binding to bilayers to see what insights vibrational spectroscopy can bring to the mechanism of its insertion and possible translocation to the other side of the membrane.
Much of the work is by attenuated total internal reflection FTIR SPECTROSCOPY (ATR-FTIRS), with the amide bands of the peptide giving structural information alongside the spectra of the disrupted lipid molecules. I also use total internal reflection RAMAN SPECTROSCOPY to acquire complementary spectra and measure the surface activity of indolicidin.
Around 90% of drug targets, including those of drugs already in use, are membrane proteins. Knowledge of their structures greatly aids drug design but their ‘oily’ (hydrophobic) properties have made crystallization for diffraction techniques very difficult and only around 50 structures were solved this way at the turn of this century and 40 more since. On top of the thousands of unsolved membrane protein structures, many non-membrane proteins and peptides interact with lipid membranes as part of their functions. Lipids are surfactant molecules and form bilayer lamellar phases on silica surfaces. Surface sensitive vibrational spectroscopy gives clues to protein secondary structure and conformation with the advantage that the protein or interacting peptide is interrogated in an environment that mimics a cell membrane, a planar lipid bilayer. In ATR-FTIRS, the substrate supporting the lipid bilayer is the internal reflection element that transmits the IR beam and the membrane-peptide assembly is directly probed by the evanescent wave resulting from total internal reflection.
Indolicidin is a 13-amino acid antimicrobial peptide which is found in bovine neutrophils. With record proportions of tryptophan – five residues – three prolines, two arginines and four positive charges at pH7, it has an unusual structure and its mode of interaction with the membrane is more important to understand than its shape alone. At the other end of the spectrum are large membrane proteins with several membrane-spanning sections, which are more complicated to understand from IR spectra but for which structural details are key in the design of drugs to interact with them.
The general aim of my research in the context of surface spectroscopy is to develop procedures for obtaining IR spectra of membrane proteins from single lipid bilayers in aqueous buffer. I am therefore exploring new methods of hydration of single lipid bilayers, to overcome the common problems associated with reduced mobility and denaturation of larger proteins on solid supports.
This project is funded by the EPSRC.
Apart from finishing my DPhil?! I like songs that tell stories, setting the world to rights and wooded, rocky streams and waterfalls. A recent interest in cars and planes on account of marrying a bloke needs redressing with knitting, reading about hope-inspiring people and ideas and taking to the roads on the mode of transport we both like, cycling. I wouldn’t have thought I could cover all that in 3 lines on the world wide intellect, but there you go.