Single-molecule spectroscopy, in combination with Förster resonance energy transfer (smFRET), is a sensitive molecular ruler that allows us to measure molecular distance distributions and dynamics on a broad timescale from picoseconds to hours, through multiparameter analysis of photon statistics. The smFRET toolbox can give us a detailed view into biomolecular structure, dynamics, and function, especially when combined with molecular simulations.

The Protein research centre has a MicroTime 200; time-resolved confocal fluorescence microscope with single-molecule detection. The instrument has 2 picosecond pulsed lasers (520 nm and 640 nm) and 4 single-photon avalanche detectors with ps resolution.

This enables a wide range of multiparameter analysis of fluorescently labelled biomolecules using photon statistics: static conformational heterogeneity can be assessed from transfer efficiency histograms, and the resulting subpopulations can be analysed independently for accurate interdye distances; ensemble and single‐molecule fluorescence anisotropy provides information about the rotational freedom of the fluorophores; subpopulation‐specific fluorescence lifetime decays yield intramolecular distance distributions; fluorescence lifetime imaging (FLIM) allows high-resolution imaging of surface-immobilized molecules or within living cells, and can be combined with FRET for increased resolution; fluorescence correlation spectroscopy (FCS) provides dynamic information over a broad range of timescales from picoseconds to seconds, including rapid sub-µs dynamics and translational diffusion, which yields hydrodynamic radii and reveals the potential presence of aggregates and association reactions; alternating excitation allows molecules without an active FRET pair to be identified.

The experiments are performed on either freely diffusing molecules, which restricts dynamic processes to the millisecond range or on surface-immobilized molecules which allows conformational or binding kinetics to be measured on a timescale from milliseconds to minutes.

X-ray crystallography is a method where protein crystals are formed following by measuring series of X-ray diffraction images from an X-ray source in order to solve the structure of the protein biomolecule.

Our protein crystallography facility has all the equipment for growing monitoring protein crystal growth.We utilize both commercial and inhouse made crystal screens in order for screen for good crystal growing conditions.

Our facility will soon be equipped with a NT-8 dropsetter (Formulatrix) which allows screening of nanoliter scale volume of proteins which is also equipped with a luiquid-cubic-phase (LCP) module designed for membrane protein crystallization.

The facility has cryoshipping equipment ready to use to send to synchrotron facilities throught EMBL. We also have access to a single-X-ray crystal instrument (Bruke D8 venture) in collaboration with the Department of Chemistry to screen for diffraction.

We have access to a 600 MHz Bruker Avance NMR spectrometer with 1H, 15N, and 13C probes for protein analysis.

We have expertise in protein isotope labelling and multidimensional NMR analysis of protein structure and dynamics.

Most of the proteins that we research are expressed in E.coli. We utilize various strategies and differents strains of E.coli to allow us to express diffucult to express proteins in soluble form.

We also have facilities to express proteins in mammalian suspension culture and insect cell lines.

Our laboratories are equipped with an Akta Pure FPLC instrument, equipped with an F9-C fraction collector which is our routine protein chromatography instrument.