Final Report Summary - SELFBIOLOGICS (Mimicry of biology supramolecular logics towards self-assembly of artificial components for life) Whether a cell adheres to the extracellular matrix, or biological signals propagate within and between cells, highly selective interactions occur between the molecular partners that materialize the process. Supramolecular chemistry studies the basic features of these interactions (knowledge) and their implementation for the design of non-natural systems (technology). This field bridges molecular chemistry and physics with biology, providing a interdisciplinary platform to understand biological structure (self-assembly) and function (recognition, reactivity and transport). Peptides that self-assemble in ordered nanostructures are a particular case of supramolecular chemistry. Peptides possess the biocompatibility and chemical diversity found in proteins, being particularly interesting for regenerative medicine and nanomedicine. Until now, peptide self-assembly systems have been studied individually. However, biological structures form in highly dense and heterogeneous molecular environments, such as the cytosol and extracellular matrix. The main scientific objective of this project is to recreate part of this complexity, by creating multi-component peptide self-assembly systems that form independent nano-assemblies in the same physical space. It has been shown in the past decade that peptides that self-assemble in fibrilar nanostructures possess a great potential in regenerative medicine applications. These include regeneration of tissues that have been classically regarded as very difficult to regenerate, such as cartilage, as well as neuronal and cardiac tissues. Furthermore, peptide self-assembly systems can be designed to create injectable materials (liquid formulations that gel upon contact with the body fluids), providing a platform for minimally invasive delivery of regenerative therapies. The medical applications for peptide self-assembly based materials foresee a huge socio-economic impact potential. The virtue of self-assembly is that the structural features (both at micro and nanoscales) do not depend on human intervention (nor external devices), but they are molecularly encoded in the peptide structure. In this way, injectable therapies can be developed, still retaining control over the fine nanostructure, as well as over the hierarchical organization over several length scales. The goal is to create artificial materials that replicate the biological features and nanostructure of the natural extracellular matrix. These materials should work as a temporary scaffold designed to signal cells, triggering them to recapitulate the tissue morphogenesis. This scaffold should be eliminated once cells take over the regeneration process. Degradation products should be non-toxic and eliminated through excretion and/or natural metabolic pathways. Alternatively, materials can break down in natural building blocks such as amino acids (as in the case of peptides), sugars and/or fatty acids, and be finally reincorporated in the newly formed tissues. Replicating the biological complexity of the extracellular matrix requires the ability to control the molecular self-assembly process, but also to create both self-sorting and co-assembling molecules. Since complexity without control is aimless, this complexity should be built over combining layers of simpler and robust self-assembly units. In order to attain this control we studied how amino acid sequences influence the dynamics of molecular exchange between self-assembled nano-objects. Since self-assembly is caused by non-covalent interactions between the molecular building blocks that compose the final supramolecular structure, it is somehow expected that molecules can be dynamically exchanged between preformed nanostructures, if not kinetically trapped. This means that molecules might be able to leave the nanostructure, become water soluble and reassemble in a different preformed nanostructure, or nucleate a new one. For instance, it has been known that cell membrane lipids, a well-known example of self-assembly in a biological context, possess the remarkable ability to migrate between biomembranes.We studied how fast is the exchange dynamics of our peptide self-assembly systems, and how the amino acid sequence affects the exchange rate. This has been undertaken using fluorescence techniques, such as Foster resonance energy transfer (FRET) and fluorescence anisotropy. Since the FRET experiment only gives information about the entire ensemble, it is not possible to take conclusions about the mechanism of the molecular exchange process. To elucidate the mechanism, it would be important to know the dyes distribution along the fibers long axis with time during the exchange. In order to determine the distribution of dyes, we have used super-resolution Stochastic Optical Reconstruction Microscopy (STORM). Understanding mechanistic and dynamic processes occurring in peptide self-assembly is an important piece of information to design more sophisticated artificial extracellular matrices. In this sense, the fundamental aspects of peptide self-assembly studied have a great potential to be readily translated into medicine.