A multi-pore model of the blood-brain barrier tight junction strands recapitulates the permeability features of wild-type and mutant claudin-5

Abstract

In the blood-brain barrier (BBB), endothelial cells are joined together by multi-protein assemblies called tight junctions (TJs), which seal the paracellular space and restrict the passage of substances. Among the proteins forming BBB TJs, Claudin-5 (Cldn5) is the most abundant one. Structural models for complexes of Cldn5 and other claudins have been proposed and assessed via experimental and computational approaches. In these models, first introduced for channel-forming, selectively permeable claudins, protomers are arranged to form pores in the paracellular space that regulate transport by electrostatic and/or steric effects arising from the pore-lining residues. With limited exceptions, however, previous computational studies focused on oligomers of only a few subunits, while extended polymeric claudin strands form TJs. Here, we employ multi-microsecond all-atom molecular dynamics and free energy (FE) calculations to study two distinct models of TJ-forming Cldn5 complexes, called multi-Pore I and multi-Pore II, each comprising sixteen protomers arranged around three adjacent pores. Free energy calculations of water and ions permeation across the pores reveal that, in both models, the passage of ions is hindered by FE barriers, which are higher than in single-pore architectures. Moreover, only the multi-Pore I structural model recapitulates the effect of the G60R variant of Cldn5, making it permeable to anions. The results provide new insights into Cldn5 structure and function and validate a structural model of BBB TJs that may be useful for the study of barrier impairment in brain diseases and for developing new therapeutic approaches.