Methodology development in biomolecular spectroscopy
Working group chair – Claus Seidel (Germany)
A wide range of spectroscopic techniques (UV, visible, near-IR, vibrational mir-IR, Raman and magnetic resonance techniques) have long been used successfully to study biological molecules with suitable chromophores in solution as well as, in a subset of systems, the crystal. The ability to correlate structure with both function and dynamics requires that these techniques be applied in a time resolved manner and, indeed, many of these spectroscopies are amenable to fast time resolved data collection provided that methods are available for initiation and repetition of the fast transient events. It is this initiation of repetitive cycles that is often the limiting factor. Where reversible photochemistry is not possible, other possible strategies are needed, both to initiate transients – for example by rapid mixing in jets or stopped-flow, and to deal with irreversible systems – for example by sample exchange methods. Long range structural constraints imposed by nanometer distance measurements are available through labelling methods in conjunction with both EPR and FRET. These are presently limited in accuracy by type of label, their mobilities and in maximum distance estimates as a result of machine sensitivities. Thus far they have also not yet been adapted to routinely perform time resolved experiments.
Working group objectives
1) To address many of these limiting factors by stimulating exchanges of information between spectroscopic instrument makers and those with expertise in new methods of transient initiation/repetition;
2) To create and update a comprehensive database of the unique instrumentation in Europe;
3) To create links between technologists and users with specific biological questions in order to provide access opportunities for applications to genuine biological problems;
4) To train new researchers in these areas through Workshops and Training Schools.
Specific goals of WG3 are:
1) To develop, and provide access to, methods of transient initiation/repetition, specifically addressing issues of improved and wider applicability of photochemistry, non-photochemical means of transient initiation and strategies to deal with irreversible systems;
2) To develop new and more rigid labels to improve distance accuracies in EPR/FRET measurements; to extract multiple distance information from multiply labelled samples
3) To improve EPR spectrometer performance including accessing RT distance measurements, novel pulse techniques and accessing longer distances as well as orientations.
4) Development of novel approaches to performing time-resolved distance measurements both in solution and ultimately in single crystals.