Complement activation by bacterial surface glycolipids: a study with planar bilayer membranes.

Abstract

Planar asymmetric glycolipid/phospholipid bilayer membranes were used as a reconstitution model of the lipid matrix of the outer membrane of Gram-negative bacteria to study complement (C) activation by various bacterial surface glycolipids with the aim of defining the C activation pathway. As glycolipids the lipopolysaccharides of Salmonella enterica serovar Minnesota R mutant strains R595 (Re LPS) and R4 (Rd2 LPS), pentaacyl lipid A from the LPS of the Escherichia coli Re mutant F515, and glycosphingolipid GSL-1 of Sphingomonas paucimobilis IAM 12576 were used. Methylester and carboxyl-reduced derivatives of GSL-1 were used to elucidate the role of the carboxyl group as common functional group of LPS and GSL-1 for C activation. The formation of lytic pores was monitored via the measurement of changes in membrane current. For all glycolipids we observed a considerable increase in membrane current soon after addition of whole human serum due to the formation of lytic pores in the membranes. Pore formation was dependent on the presence of C9, indicating that the observed current changes were due to C activation. We found that in our reconstitution system of the outer membrane lipid A, Re LPS, and Rd2 LPS activated the classical pathway, the activation being independent of specific anti-LPS antibodies. In contrast, GSL-1 and the methylester derivative of GSL-1 activated the alternative pathway even at the low serum concentrations used in this study (about 0.2% v/v). Interestingly, the carboxyl reduced GSL-1 activated the classical pathway.