Abstract
Complexes containing lipopolysaccharide (LPS) and three outer membrane proteins (OMPs) are released by gram-negative bacteria incubated in human serum and into the circulation in an experimental model of sepsis. The same OMPs are bound by immunoglobulin G (IgG) in the cross-protective antiserum raised to Escherichia coli J5 (anti-J5 IgG). This study was performed to identify the three OMPs. The 35-kDa OMP was identified as outer membrane protein A (OmpA) by immunoblotting studies using OmpA-deficient bacteria and recombinant OmpA protein. The 18-kDa OMP was identified as peptidoglycan-associated lipoprotein (PAL) based on peptide sequences from the purified protein and immunoblotting studies using PAL-deficient bacteria. The 5- to 9-kDa OMP was identified as murein lipoprotein (MLP) based on immunoblotting studies using MLP-deficient bacteria. The studies identify the OMPs released into human serum and into the circulation in an experimental model of sepsis as OmpA, PAL, and MLP.
Highlights
Bacterial cell wall components released into the bloodstream are believed to be important in the pathogenesis of gram-negative sepsis
We have demonstrated that immunoglobulin G (IgG) in antiserum raised to heatkilled E. coli J5 (J5 antiserum) binds to the same three gramnegative bacterial outer membrane proteins (OMPs) that are released into serum in the OMP-LPS complexes described above [30]
These results indicate that the 35-kDa OMP is outer membrane protein A (OmpA) and that 2D3 is a monoclonal anti-OmpA IgG
Summary
Bacterial cell wall components released into the bloodstream are believed to be important in the pathogenesis of gram-negative sepsis. We have affinity purified complexes containing LPS and at least three OMPs, with estimated molecular masses of 35, 18, and 5 to 9 kDa, from filtrates of normal human serum incubated with Escherichia coli bacteria, using O-chain-specific anti-LPS IgG [29, 30]. At least one OMP, with an estimated molecular mass of 18 kDa, is released from bacteria separately from the OMP-LPS complexes and in a form that is selectively affinity purified from human serum and septic rat plasma by IgG in J5 antiserum [29]. This study was performed to identify the 35-, 18-, and 5- to 9-kDa OMPs that are released in vitro into human serum [30] and in vivo into the circulation in experimental gram-negative sepsis [29] and are bound by IgG in J5 antiserum
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