The role of internal urease in acid resistance of Helicobacter pylori

Background & Aims: The relative role of internal urease for acid protection of Helicobacter pylori is unknown, The aim of this study was to determine the comparative importance of internal and external urease under acidic conditions, Methods: The pH optimum and measured Michaelis constant for urea of external urease and urease in intact bacteria at different medium pH (pH(out)) were measured using (CO2)-C-14 release from C-14-urea, The effect of urea on membrane potential and bacterial cytoplasmic pH was measured at different fixed pH(out). S-35-methionine labeling and sodium dodecyl sulfate-polyacrylamide gel electrophoresis of labeled proteins in the organism and medium measured protein synthesis at different pH(out) and mechanisms of urease externalization. Results: External urease had activity between pH 5.0 and 8.5 and internal urease between pH(out) 2.5 and 6.5, and its Michaelis constant at pH(out) 2.5 was 300 mmol/L but at pH(out) 4.5 was 0.5 mmol/L, similar to free urease, The addition of 5 mmol/L urea to bacteria at fixed pH(out) from 3.0 to 6.0 elevated potential to about -105 mV and periplasmic pH to about pH 6.2, Protein synthesis occurred mainly between pH 6.5 and 8.0, and urease activity resulted in increased protein synthesis at acidic pH, The labeling pattern of intrabacterial and released protein was similar. Conclusions: Intracellular urease activity is regulated by external pH, defends against gastric acidity by increasing periplasmic pH and membrane potential, and stimulates protein synthesis at acidic pH. External urease is produced mostly by cell lysis.