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The amount of F 1 Fo-ATPase in Enterococcus hirae (formerly Streptococcus faecalis) increases when the cytoplasmic pH is lowered below 7.6, and protons are extruded to maintain the cytoplasmic pH at around 7.6. In the present study, we found that the transcriptional activity of the F 1 Fo-ATPase operon was not regulated by pH. The synthesis of F 1 subunits was increased 1.65^0.12-fold by the acidification of medium from pH 8.0 to pH 5.3. Western-blot analysis showed that there were F 1 subunits in the cytoplasm, and the number of alpha plus beta subunits in the cytoplasm was 50% of the total number of the subunits in cells growing at pH 8.0. This decreased to 22% after shifting the medium pH to 5.3, with a concomitant 5.1-fold increase in the level of membrane-bound F 1 Fo-ATPase. The cytoplasmic F 1 subunits were shown to be degraded, and Fo subunits not assembled into the intact F 1 Fo complex were suggested to be digested. These data suggest that regulation of the enzyme level of F 1 FoATPase by the intracellular pH takes place mainly at the step of enzyme assembly from its subunits.Keywords: Enterococcus hirae; F 1 Fo-ATPase; assembly; pH regulation.Since the first demonstration in the anaerobic bacterium Enterococcus hirae (formerly Streptococcus faecalis) that cytoplasmic pH is regulated by proton extrusion mediated by a proton-translocating ATPase [1], many reports have shown that the H + -ATPase is involved in pH homeostasis in prokaryotes and eukaryotes [2,3]. In eukaryotes, a V-type H + -ATPase regulates the internal pH, while the regulator of the enterococcal cytoplasmic pH is an F-type enzyme. Because V-type and F-type H + -ATPases are very similar in structure and catalytic properties, the basic mechanism for pH regulation should be the same.In prokaryotes, it was already known by 1984 that the amount of the enterococcal F 1 Fo-ATPase was increased by lowering the cytoplasmic pH [4,5]. Both experimental [6] and theoretical [7] data have shown that internal pH homeostasis is maintained by changes in the amount of this enzyme. However, how the enzyme level is regulated is still unclear.Smith et al.[8] have shown no stimulation of the transcription of the H + -ATPase operon at low pH in S. mutans, in which the enzyme level is regulated by pH. In the present study, we found that the transcriptional activity of the F 1 Fo-ATPase operon was not affected by pH in E. hirae, while its translation was stimulated at low pH, but the stimulation was too low to account for the increase in the level of membrane-bound F 1 FoATPase. Our present data suggest that the enzyme level is regulated mainly during the step in which the enzyme is assembled from its subunits. MATERIALS AND METHODS Bacterial strains, plasmids and culture mediaEnterococcus hirae was grown at 378C in either medium 2KTY [1] or the following media without shaking. Medium A was a synthetic medium described previously [9] with the following modifications. Solution A contained 50 mm ammonium sulfate, 8 mm magnesium sulfate, 0.5 mm manganese sulfate, 0...

The chemical composition and immunobiological activities in vivo and in vitro of sodium dodecyl sulphate extracts (SDS-SE) derived from periodontopathic bacteria (three strains of Actinobacillus actinomycetemcomitans, two strains of Bacteroides gingivalis, and one strain of Fusobacterium nucleatum) were investigated. The main components of SDS-SE were protein and lipid, with negligible amounts of peptidoglycan and lipopolysaccharide. Immunopotentiating activity was detected in both delayed-type hypersensitivity and antibody formation against the elicitation of a protein antigen with the SDS-SE preparations of A. actinomycetemcomitans ATCC 29524 and B. gingivalis 381 and 1021. On the other hand the SDS-SE of A. actinomycetemcomitans ATCC 29522 enhanced only the induction of a delayed-type hypersensitivity response. All the SDS-SE preparations had mitogenic activity to splenocytes from BALB/c nu/nu, C3H/HeN and C3H/HeJ mice. Migration-stimulating activity for human peripheral blood monocytes was detected especially in the SDS-SE preparations of A. actinomycetemcomitans ATCC 29524 and Y4. All of the SDS-SE samples inhibited [3H]thymidine uptake in human gingival fibroblasts and caused degradation of the cells. The results suggest that the cell membrane components extractable with sodium dodecyl sulphate from periodontopathic bacteria are involved in the pathogenesis of periodontal disease.

Immunobiological activities of sodium dodecylsulfatesoluble fractions derived from the cell envelopes of oral gram-negative, anaerobic rods