Arginine 66 Residue of Fur is Required for the Regulatory Function of this Protein in the Acid Adaptation Mechanism of Helicobacter pylori

Abstract

Background: Helicobacter pylori colonizes the gastric mucosa and must survive the acid pH of that environment. Like other enteric bacterial pathogens, including Salmonella enterica, H. pylori develops an acid tolerance response that is dependent on the function of the transcriptional regulator protein Fur.

Objective: To explore by site-directed mutagenesis whether two particular amino acid residues in the amino acid sequence of the H. pylori Fur protein, arginine 66 and histidine 99, are involved in the acid response mechanism in this bacterium.

Materials and Methods: Complementation assays in Escherichia coli H1780 ( fur null mutant) both with plasmids carrying the H. pylori fur gene bearing substitution mutations R66A or H99A or R66A/H99A and with the H. pylori Fur-R66A mutant were conducted. Wild-type and mutated Fur proteins from H. pylori were assayed by using the fiu::lacZ reporter gene in the E. coli H1780 heterologous system at various pH and iron concentrations.

Results: Both bacterial growth and repression of the reporter gene were impaired under acid conditions in E. coli H1780 complemented with pUC19-fur-R66A. Also, in the H. pylori Fur-R66 strain bacterial growth and speA gene expression were impaired under acid conditions.

Conclusions: Arginine 66 but not histidine 99 in H. pylori Fur is required for the regulatory function of the Fur protein in the acid adaptation mechanism of the bacterium.