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PEPR - A Centre for Pulse EPR will be coming soon!

PEPR - Roessler  lab We have recently secured a £2.3 M strategic equipment award from the EPSRC to build a Centre for Pulse EPR spectroscopy (PEPR) that will enable detailed insight into the structure and dynamics of paramagnetic compounds. Besides encompassing state-of-the-art pulse EPR instrumentation at X- and Q-band frequencies, coupled to photoexcitation, we will work in collaboration with John Morton’s group at UCL to push the detection limit of spins and in pulse EPR investigations of anisotropic spin systems such as respiratory and photosynthetic complex I. Moreover, with PEPR we will further develop our combination of film electrochemistry and EPR spectroscopy, to enable new breakthroughs in the characterisation of redox processes.

Watch this space for further details, including PEPR’s facility manager position and PhD studentships!

Research overview

Oxidation-reduction (redox) reactions underpin innumerable chemical reactions - and much of the chemistry of life! We are fascinated by how oxidation-state changes govern respiration and photosynthesis and how nature has fine-tuned the redox properties of its many intricate molecular machines. Redox reactions often involve transition metal ions and our group is investigating the properties, structure and bonding of centres with unpaired electrons in complex biological machines.

Many redox reactions proceed via radical intermediates and these are, perhaps luckily, often located in mechanistically key locations. We use electron paramagnetic resonance (EPR) spectroscopy as a powerful method for obtaining detailed information on the structure and bonding in these ubiquitous spin centres. Electrochemistry on the other hand, in particular film electrochemistry, provides insight into the reactions carried out by molecular machines. These (bio)physical methods are supplemented by biochemical methods.

Method Development

A new spectroelectrochemical method 

PFE-EPR setup

The combination of electron paramagnetic resonance (EPR) spectroscopy with protein film electrochemistry (PFE) provides a novel platform for the investigation of redox-active species including metalloproteins. The development of PFE-EPR enables the detection of paramagnetic species with direct and accurate potential control, providing new mechanistic insight to redox-based processes in biomolecules, including catalysis. Current EPR spectroelectrochemical methods to study proteins are based on generating radical species in solution and therefore limited by diffusion (precluding accurante potnetial control) and do not enable catalytic investivations. We exploit the advantages of PFE, i.e. 'wiring' electrons directly to the working electrode surface, to eliminate diffusion limitations. 
We have developed a generally applicable EPR spectroelectrochemical method to study biomolecules irrespective of their size: PFE-EPR. We prove the feasibility of a combined PFE-EPR setup using small paramagnetic molecules and proteins, 4-amino TEMPO and Cu-Zn superoxide dismutase (16 kDa dimer) and demonstrate that 3-dimensional (cylindrical) hierarchical indium tin oxide (ITO) structures are suitable working electrode materials, offering good conductivity and large surface area for protein immobilisation. The resulting EPR spectrum of SOD/TEMPO immobilised on meso-ITO electrode not only indicates successful reduction and detection of the paramagnetic species but also signifies the direct, accurate and fast potential control offered by the PFE-EPR setup. We are now developing PFE-EPR further to use this new technique to investigate the mechanism of large metalloproteins such as respiratory complex I. 

Project members and collaborators

  • Kaltum Abdiaziz (PhD student, QMUL & ICL)
  • Adam Sills (PhD student, ICL)
  • Dr. Maryam Seif Eddine (PDRA, ICL)
  • Dr. Sam Cobb (PDRA, Cambridge)
  • Pr. Erwin Reisner (PI, Cambridge)

Respiratory Complex I

Complex I (NADH:ubiquinone oxidoreductase)

Respiratory CxI

Complex I – the entry point of electrons into the respiratory chain of all higher organisms – is the last of the respiratory enzymes whose mechanism remains unknown. In mitochondria, complex I oxidises NADH from sugars and fats, reduces ubiquinone and transports protons across the inner mitochondrial membrane, thereby contributing to the proton motive force that enables ATP-synthase to make ATP.

We are working on elucidating some of the key elements of the mechanism of complex I using advanced EPR methods, in particular we are fascinated how the redox energy built up through electron transfer is used to drive proton translocation. Given the large size of the enzyme (the size of the mitochondrial enzyme is ca. 1 MDa), many biophysical methods routinely applied to study protein cannot be used (e.g. NMR). We are exploiting the fact that EPR spectroscopy is blind to all the many paired electrons in the complex, and that unpaired electrons are situated in mechanistically key locations. For instance, many of the iron-sulfer clusters ([2Fe-2S] and [4Fe-4S] clusters) become EPR-visible when reduced. We have recently divised a potentiometric method that enables us to adjust very small amounts of protein to very precise reduction potentials - an important prerequisite for studying these redox-active proteins spectroscopically. This has enabled us, in conjunction with pulse EPR spectroscopy and site-directly mutagenesis, to investigate the role of Fe-S cluster N2

Project members and collaborators

  • Dr. Maryam Seif Eddine (PDRA, ICL)
  • Kaltum Abdiaziz (PhD student, QMUL)
  • Adam Sills (PhD student, ICL)
  • Dr John Wright (PDRA, Medical Research Council, Mitochondrial Biology Unit, Cambridge)
  • Dr Judy Hirst (PI, Medical Research Council, Mitochondrial Biology Unit, Cambridge)

Photosynthetic Complex I

The NAD(P)H dehydrogenase 'like' complex (NDH) 

Photosynthetic Complex I

We have teamed up with Dr Guy Hanke to investigate the fascinating molecular machine NDH, in many ways related to the mitochondrial enzyme (complex I, above), present in plants and cyanobacteria. Our goal is to unravel the mechanism of NDH, and ultimately to exploit this knowledge in order to increase stress tolerance in plants (in collaboration with Prof. Nestor Carrillo, Rosario, Argentina). Increasing crop yields is an especially important challenge facing the developing world, but may affect us all one day given the ever rising world population. Research on NDH has thus far focused on genetic investigations and very little is known about the function of NDH, and no biophysical studies are reported. Using our experience with the mitochondrial enzyme, we are using EPR spectroscopy and protein film electrochemistry to investigate the co-factors in and the electron donors to NDH. Moreover, the knowledge gained through NDH is also likely relevant for understanding the mechanism of complex I. 

See Dr Hanke's webpage for more details on the biological context. 

Project members and collaborators

  • Gemma McGuire (PhD student, QMUL & ICL)
  • Katherina Richardson (PhD student, LIDo program, QMUL & ICL)
  • Dr Guy Hanke (PI, School of Biological and chemical Sciences, QMUL)

Electron paramagnetic resonance

ELectron Paramagnetic Resonance  


Electron paramagnetic resonance (EPR) spectroscopy is the study of materials or molecules with unpaired electrons. The technique makes use of the intrinsic angular momentum of an electron – its ‘spin’ – by placing the paramagnetic material into a magnetic field and applying microwave radiation. 

For a given molecule, such as the fictive molecule in the figure below, an EPR field sweep (traditionally performed by continuous wave EPR) provides us with information on the type of chemical centre the unpaired electron resides on. However, spectral and time resolution is limited and exploiting the full potential of EPR spectroscopy requires pulsed methods. These allow the separation of different types of interactions.

EPR applications

Fig. 1: Illustration of the use of EPR for structural analysis. The position of the unpaired electron is indicated by the full black circle in the fictive molecule. The coloured rings and corresponding schematic spectra indicate which parts of the molecule can approximately be studied with a particular EPR technique. Figure adapted from C. Calle et al.Chimia,2001, 55, 763.

Hyperfine techniques such as ENDOR (electron nuclear double resonance), ESEEM (electron spin-echo envelop modulation) and HYSCORE (hyperfine sublevel correlation) spectroscopy yield information on interactions with more distant nuclei and spectral resolution can be improved by orders of magnitude. We can thus learn about distances between electron and nuclear spins, the spin density distribution (the type of bonding), and electric field gradients.

Double electron-electron resonance (DEER), or synonymously pulse electron-electron double resonance (PELDOR) spectroscopy, has been very widely applied to many different types of systems, especially in biology, to determine the distance between two unpaired electrons.

Whilst the majority of EPR experiments are carried out at the X-band frequency, going to higher field and frequencies (e.g. Q- and/or W-band) can be extremely useful in detangling complicated spectra, by separating field dependent from field-independent components and thereby increasing spectral resolution.


We currently have an X-band continous-wave EPR spectrometer at the MSRH. We also use the Imperial SPIN-Lab in South Kensington and the pulse EPR spectrometers at UCL (group of Prof. John Morton).