Biochemical and immunological properties of Coxiella burnetii cell wall and peptidoglycan-protein complex fractions

Abstract

Coxiella burnetii morphological cell types were fractionated into large-cell variant cell walls, two fractions of small-cell variant cell walls, and one fraction of small-cell variant whole cells. Based on the contents of peptidoglycan (PG)-constituents and the yields of the sodium dodecyl sulfate-insoluble PG-protein complex (PG-PC) from cell walls, the fraction of large-cell variant cell walls contained significantly less PG than did the fraction of small-cell variant cell walls. The yields of PG-PC from the fractions of large-cell variant cell walls and small-cell variant cell walls were 2 and 32%, respectively. These results indicated that the PG of the largecell variant cell walls may be partially digested by PG-lytic enzymes or incompletely synthesized, whereas the small-cell variant cell walls appeared to have intact PG. Proteins associated with PG-PC were resistant to proteolysis by trypsin, protease VI, and proteinase K. Saturated and unsaturated fatty acids were detected in whole cells and cell walls but not in PG-PC, which contained a 3-deoxy-D-mannooctulosonic acid-like component that is also present in phase I lipopolysaccharide. Immunogenicity of the fractions was tested by measuring the temporal sequence of phase II and phase I antibody responses in vaccinated rabbits. Both phase II and phase I antibody responses were demonstrated with all fractions except the sodium dodecyl sulfate supernatant of the small-cell variant cell walls, whereas PG-PC elicited a pure phase II antibody response up to 29 days postvaccination. The immunogenicity of these fractions may reflect a quantitative difference in antigen concentration or may be due to a qualitative difference in phase II and I determinants.

Sensitivity of Coxiella burnetii peptidoglycan to lysozyme hydrolysis and correlation of sacculus rigidity with peptidoglycan-associated proteins

“…Based on the results of our preceding paper, the PG isolated by Schramek et al (21) may correspond to the PG-PC which was purified from SCV-CW (4). This PG-PC was resistant to proteolysis with some proteases (4). In the present study, PG-associated proteins also were resistant to solubilization with SDS and 2-ME (Table 1).…”

“…Organism. All procedures with phase I C. burnetii, Ohio isolate, were carried out as described previously (4).…”

“…Analytical methods. Quantitative and qualitative analyses of amino acids, amino sugars, proteins, neutral sugars, reducing groups, 3-deoxy-D-mannooctulosonic acid (KDO), and fatty acids were performed as described previously (2,4).…”

Steric hindrance of antibody binding to surface proteins of Coxiella burnetti by phase I lipopolysaccharide

“…The exposure of shared phase I and II surface proteins was demonstrated by direct binding of 1251I-anti-phase II protein antibodies to TCA-extracted phase I cells, as well as the increased reactivity of these extracted phase I cells with anti-phase II antibodies in immunofluorescence assays and immunoelectron microscopy studies. Similarly, Williams et al (30) described a monoclonal antibody that recognized a 29.5-kilodalton protein of phase II but not native phase I C. burnetii and subsequently interpreted this as indicating that phase II specificity resided in that protein (2). Interestingly though, the protein could be exposed on phase I cells by chloroform-methanol extraction.…”

“…Phase-dependent differences in surface proteins or antigens have been described (31). A study with monoclonal antibodies (30) has led to the opinion that phase II specificity resides in a 29.5-kilodalton surface protein (2). However, we have reported (12) that the protein components of phase I and II cells are for the most part shared and that the unique phase-dependent antigens are the LPSs.…”