A combination of proteomics, genetics, and physiology provides insights into the acid-tolerance phenotype of Pseudomonas pergaminensis F77

Acid tolerance is crucial for the effective and persistent mineral weathering by acid-producing bacteria. Here, the molecular basis of the acid tolerance of mineral-weathering Pseudomonas pergaminensis F77 was identified using proteomics analysis of the strain under acid stress. Then, the acid tolerance of strain F77 and its mutants with deletion of the acid tolerance-related genes orf03767, mcp, resR, nueR, yegD, and fxsA, which are involved in the two-component systems, DNA repair, nucleotide binding, and membrane parts, were compared. Finally, the acid tolerance-related physiological mechanisms of strain F77 and its mutants F77 Delta nueR and F77 Delta resR under acidic conditions were characterized. The significantly upregulated proteins in the acid-adapted and acid-challenged strain F77 included the proteins involved in metabolic pathways associated with ATPase, membrane compo-nents, organic acid transmembrane transporters, response to stimulus, nucleotide binding, ABC transporters, and two-component systems. The cell numbers decreased by 24-100% at pH <= 4.50, while the membrane fluidity increased by 22-61% at pH <= 5.50 for the mutants F77 Delta nueR and F77 Delta resR, compared with that of strain F77. The intracellular H+-ATPase activities decreased by 29-33% for the mutant F77 Delta nueR at pH <= 4.50% and 33-79% for the mutant F77 Delta resR at all tested pHs (pH <= 7.00); meanwhile, the ratios of intracellular NAD+/ NADH decreased by 71-91% for the mutant F77 Delta resR at all tested pHs (pH <= 7.00), compared with that of strain F77. Furthermore, the intracellular putrescine concentrations were reduced by 40-70% for the mutant F77 Delta resR at all tested pHs (pH <= 7.00) compared with that of strain F77. Our findings suggested that multiple proteins and metabolic pathways were associated with bacterial acid tolerance and revealed that nueR and resR were involved in acid tolerance based on their modulation of multiple acid tolerance-related physiological functions in strain F77.