Mechanobiology - chemical origin of membrane mechanical resistance and force-dependent signaling.

The cell membrane is a highly complex designed material with remarkable physicochemical properties; comprised mainly of lipid moieties, it is capable of self-assembling, changing morphology, housing a range of distinct proteins, and withstanding electrical, chemical and mechanical perturbations. All of these fundamental cellular functions occurring within a 5nm thick film is an astonishing feat of engineering, made possible due to the interplay of a variety of intermolecular forces. Elucidating how the interactions within the chemically distinct partners influence the nanomechanical properties of the membrane is essential to gain a comprehensive understanding of a wide-variety of both force-triggered and force-sensing mechanisms that dictate essential cellular processes.

Large-scale modelling of the divergent spectrin repeats in nesprins: giant modular proteins.

Nesprin-1 and nesprin-2 are nuclear envelope (NE) proteins characterized by a common structure of an SR (spectrin repeat) rod domain and a C-terminal transmembrane KASH [Klarsicht-ANC-Syne-homology] domain and display N-terminal actin-binding CH (calponin homology) domains. Mutations in these proteins have been described in Emery-Dreifuss muscular dystrophy and attributed to disruptions of interactions at the NE with nesprins binding partners, lamin A/C and emerin. Evolutionary analysis of the rod domains of the nesprins has shown that they are almost entirely composed of unbroken SR-like structures. We present a bioinformatical approach to accurate definition of the boundaries of each SR by comparison with canonical SR structures, allowing for a large-scale homology modelling of the 74 nesprin-1 and 56 nesprin-2 SRs. The exposed and evolutionary conserved residues identify important pbs for protein-protein interactions that can guide tailored binding experiments. Most importantly, the bioinformatics analyses and the 3D models have been central to the design of selected constructs for protein expression. 1D NMR and CD spectra have been performed of the expressed SRs, showing a folded, stable, high content α-helical structure, typical of SRs. Molecular Dynamics simulations have been performed to study the structural and elastic properties of consecutive SRs, revealing insights in the mechanical properties adopted by these modules in the cell.