Molecular dynamics simulations of tight junction proteins

Fitsiou, Eleni and Anwar, Jamshed (2020) Molecular dynamics simulations of tight junction proteins. PhD thesis, Lancaster University.

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Abstract

Tight junctions (TJs) are specialised cell-cell structures that serve primarily as a barrier to molecular transport through the intercellular space between the cells. The claudin family of proteins are the main structural and functional components of the TJ strands that circumscribe the cells. The detailed molecular organisation at the TJs is not entirely resolved, being relatively inaccessible by current experimental methods. Here, we have employed molecular dynamics simulations using both atomistic and coarse-grained models to investigate the TJ structure formed by claudin-1 using self-assembly coupled with free energy calculations and enhanced sampling techniques. A feature of the studies is that the self-assembly simulations have been carried out using atomistic detail (a first) by simulating only the extracellular domains of claudin-1 in an implied membrane. The results show that the cis-interaction can occur in the absence of trans-interacting partners and that a claudin dimer is the smallest stable unit. The dimers further form higher-order aggregates with a plethora of interacting dimeric interfaces. The transinteraction of claudins resulted in a compact structure with a minimal pore size confirming the barrier properties of claudin-1. The simulations also enabled the identification of the key regions of the claudin responsible for the trans-interaction, with the identified important amino acids being in agreement with experimental studies. The role of the lipid environment, with a focus on the skin lipids in the stratum granulosum, was also investigated, along with the effect of single-point mutations in claudin-1. The single-point mutation studies were consistent with experimental results. The simulation studies have enhanced our understanding of the assembly and structure of claudin-1 TJs, a notable finding being that kinetic locking is likely to be important in determining the TJ structure. The single-point mutation studies suggest that simulations could serve as screens towards defining potential gene-therapy strategies.