title: Complement resistance in Borrelia burgdorferi strain 297: outer membrane proteins prevent MAC formation at lysis susceptible sites.
abstract: Two variants of Borrelia burgdorferi strain 297, complement-resistant wild-type (WT297) and complement-sensitive mutant (MUT297), were used as a model to study the mechanism of resistance to the alternative complement pathway in this organism. No difference in the quantity of membrane attack complex (MAC) deposition on WT297 and MUT297 was observed after 2 h incubation with normal human serum (NHS), at which time 4% of WT297 and 95% of MUT297 were killed. The polymerization of C9 bound to WT297 and MUT297 was demonstrated by immunoblotting using an anti-C9 polyclonal antibody. Immunofluorescence and thin-section immunoelectron microscopy showed MAC to be diffusely distributed on the outer membrane of both variants. Furthermore, MAC appeared to be tightly bound to the surface of both variants as demonstrated by elution studies. Protease treatment rendered WT297 susceptible to killing by NHS, suggesting that outer membrane proteins may be associated with complement resistance of WT297. One- and two-dimensional gel electrophoreses showed that proteins of 20 and 30 kDa, and 66 kDa were present in WT297 but were absent or sparse in trypsin-treated WT297 and MUT297. Interestingly, immunoblotting using a polyclonal antibody against C3 showed that C3 fragments appeared to bind different acceptors on WT297 than on trypsin-treated WT297, or MUT297. Therefore, the binding of C3 fragments to acceptors on WT297, in contrast to MUT297, may not direct the formation of the MAC to lysis-susceptible sites on the surface of the bacterium, resulting in the complement resistance of WT297.
authors.affiliation: Department of Microbiology and Immunology, Georgetown University School of Medicine, Washington, DC, USA.
authors.forename: K
authors.initials: K
authors.lastname: Patarakul
authors.forename: M F
authors.initials: MF
authors.lastname: Cole
authors.forename: C A
authors.initials: CA
authors.lastname: Hughes
mesh_terms: D000911:Antibodies, Monoclonal; D001425:Bacterial Outer Membrane Proteins / Q000276:immunology* / Q000502:physiology; D015153:Blotting, Western; D015748:Borrelia burgdorferi Group / Q000254:growth & development / Q000276:immunology* / Q000472:pathogenicity; D003176:Complement C3 / Q000276:immunology; D003186:Complement C9 / Q000276:immunology* / Q000502:physiology; D015938:Complement Membrane Attack Complex / Q000276:immunology* / Q000502:physiology; D003170:Complement Pathway, Alternative / Q000502:physiology; D015180:Electrophoresis, Gel, Two-Dimensional; D004591:Electrophoresis, Polyacrylamide Gel; D004797:Enzyme-Linked Immunosorbent Assay; D006801:Humans; D008193:Lyme Disease / Q000382:microbiology; D008856:Microscopy, Fluorescence; D016253:Microscopy, Immunoelectron; D014357:Trypsin / Q000737:chemistry
chemical_list: D000911:Antibodies, Monoclonal; D001425:Bacterial Outer Membrane Proteins; D003176:Complement C3; D003186:Complement C9; D015938:Complement Membrane Attack Complex; D014357:Trypsin