Final Activity Report Summary - PROTEIN BIOPHYSICS (Experimental and Computational Investigations of Folding, Structure, Function and Aggregation) The EXT-042334 project targeted an initiative towards developing Molecular Biology to the same quantitative level of Physics and Chemistry that involved the transfer of a multidisciplinary group in protein biophysics from the US to Europe. The main objective of the project was to exploit this move to nucleate the formation of an International Center of Excellence in Quantitative and Physical Biology in Spain that could compete with the research on this area being carried out in the US. Scientifically, the project focused on the investigation of the conformational-functional behaviour of proteins using a multidisciplinary approach that borrows concepts and methodologies from condensed matter physics, physical chemistry, molecular spectroscopy, computer science, protein chemistry, and molecular and cell biology. The scientific program was divided into four areas: 1) in-depth exploration of protein folding ensembles; 2) structure prediction and design through the calculation of folding free energy surfaces; 3) understanding downhill folding and the molecular rheostat hypothesis; 4) new concepts and approaches in protein aggregation. The original goal of the project has been fully achieved so that after the four years of the project there remains a fully operational group in protein biophysics at the host institution with a highly international member composition. In addition, a set of top of the line biophysical facilities to study protein structure and dynamics has been built, including ultrafast laser spectroscopy, single-molecule spectroscopy, atomic force microscopy, nuclear magnetic resonance and high-performance computing. The research activities of the team have resulted in a large number of publications in high profile journals and the frequent invitation of the team leader to provide oral presentations at conferences and workshops as well as deliver lectures in several academic institutions worldwide, having thus established the group as a major international player in the protein biophysics field. The members of the team have been trained in modern multidisciplinary biophysical approaches, learning molecular biology and protein biochemistry together with skills in performing quantitative experiments using a variety of sophisticated spectroscopic techniques to obtain structural, thermodynamic and kinetic information about proteins, and computing methods for multivariate data analysis. Among the most interesting scientific results of the team, we can highlight the first detailed analysis of the nature of the free energy barriers to protein folding, including methods to measure the height and energetic properties of folding barriers, experimental confirmation of theoretical predictions, and the development of methods to study the structural and energetic properties of equilibrium folding ensembles. This research has been pivotal in changing the protein folding paradigm. Other exciting results of the team have been the development of methods for structure prediction and de novo design based on the analysis of folding free energy surfaces. Important advances have also been made in testing the molecular rheostat hypothesis (namely that downhill folding proteins can exploit their inherently rich conformational behaviour to gradually modulate biological functions mimicking the mode of action of an electric rheostat), the development of quantitative methods for the experimental study of protein aggregation and for the efficient refolding of recombinant proteins from inclusion bodies (which has been the base for launching a spinoff company dedicated to the exploitation of such technology), and the extension of the downhill folding concept to engineer protein-based nanosensors with very broad sensitivity range. Finally, the added value towards the internationalisation and increase in visibility of the host institution has been enormous.
