The Effect of Media, Conditioned by Serratia proteamaculans and Enriched with ShlA Toxin, on the Permeability of Lipid Bilayers, Mimicking the Membranes of Epithelial Cells HeLa and Caco-2

Abstract— In this work we investigated the pore-forming activity of ShlA toxin from Serratia proteamaculans in lipid bilayers modelling the membranes of epithelial cells HeLa and Caco-2 and assessed the role of CaCl2 in its modification. It was found that the addition of conditioned S. proteamaculans media enriched with ShlA toxin to planar lipid bilayers formed from 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine (POPC)/sphingomyelin (SM)/cholesterol (CHOL) (33/33/33 mol %) or POPC/1-palmitoyl-2-oleoyl-sn-glycero-3-phosphoethanolamine (POPE)/1-palmitoyl-2-oleoyl-sn-glycero-3-phospho-L-serine (POPS)/SM/CHOL (30/30/5/10/25 mol %) leads to the formation of ion-permeable pores. The mean amplitude of currents flowing through single ShlA channels formed in POPC/POPE/POPS/SM/CHOL membranes was smaller than in bilayers of POPC/SM/CHOL. The current-voltage characteristics of channels formed by ShlA from S. proteamaculans in bilayers of both compositions were nonlinear. The application of media conditioned by S. proteamaculans did not increase the permeability of POPC/SM/CHOL and POPC/POPE/POPS/SM/CHOL vesicles for the fluorescent marker calcein. The addition of 10 mM CaCl2 potentiated the pore-forming activity of ShlA from S. proteamaculans in planar lipid bilayers, which manifested itself in the emergence of substates with higher conductivity. The joint addition of ShlA-enriched media conditioned by S. proteamaculans and 10 mM CaCl2 increased the permeability of POPC/POPE/POPS/SM/CHOL vesicles for calcein, but not of liposomes formed from POPC/SM/CHOL. In addition, it was demonstrated that in the presence of 10 mM CaCl2, there was a significant increase in the invasion of S. proteamaculans into Caco-2 cells, which could be mediated by an increase in the membrane activity of the ShlA toxin. The results obtained expand our understanding of the composition of the media conditioned by S. proteamaculans and the mechanisms of molecular action of the ShlA toxin on epithelial cell membranes.