Development of New Structural Characterisation Tools for Extracellular Matrix

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The project will develop new structural characterisation tools to study the molecular structure of extracellular matrix and how it is altered by non-enzymatic chemistry and cellular activity. This will include developing spectral editing tools in Dynamic Nuclear Polarisation (DNP) NMR to be able to extract NMR spectra of specific regions of the extracellular matrix from samples of intact, ex vivo matrix. For instance, we will utilise the fact that key cell adhesion proteins contact the extracellular matrix through metal-mediated binding, where the native metal ion can be replaced by a paramagnetic ion, e.g. Ca2+ can be replaced by Co2+ in integrin binding sites. Paramagnetic ions will quench the DNP effect in their locality, resulting in a DNP NMR spectrum in which NMR signals from the integrin binding site are absent. Subtraction of this paramagnetic-edited spectrum from one collected on the sample containing diamagnetic metal ions instead of paramagnetic gives an NMR spectrum of the integrin binding sites. In a second part of the project we will develop methods to measure NMR spectra on extracellular matrix whilst the matrix is under mechanical stress, with the aim of following changes in molecular conformation during that process.Overall, her project will deliver new tools to enable novel insight into biological processes highly relevant to ageing and degenerative diseases.