Bacterioferritin A Modulates Catalase A (KatA) Activity and Resistance to Hydrogen Peroxide in Pseudomonas aeruginosa

摘要

We have cloned a 3.6-kb genomic DNA fragment from Pseudomonas aeruginosa harboring the rpoA, rplQ,katA, and bfrA genes. These loci are predicted to encode, respectively, (i) the α subunit of RNA polymerase; (ii) the L17 ribosomal protein; (iii) the major catalase, KatA; and (iv) one of two iron storage proteins called bacterioferritin A (BfrA; cytochrome b ₁ or b ₅₅₇). Our goal was to determine the contributions of KatA and BfrA to the resistance of P. aeruginosa to hydrogen peroxide (H₂O₂). When provided on a multicopy plasmid, the P. aeruginosa katA gene complemented a catalase-deficient strain of Escherichia coli. ThekatA gene was found to contain two translational start codons encoding a heteromultimer of ～160 to 170 kDa and having an apparent K_m for H₂O₂ of 44.7 mM. Isogenic katA and bfrA mutants were hypersusceptible to H₂O₂, while a katA bfrA double mutant demonstrated the greatest sensitivity. ThekatA and katA bfrA mutants possessed no detectable catalase activity. Interestingly, a bfrA mutant expressed only ～47% the KatA activity of wild-type organisms, despite possessing wild-type katA transcription and translation. Plasmids harboring bfrA genes encoding BfrA altered at critical amino acids essential for ferroxidase activity could not restore wild-type catalase activity in the bfrAmutant. RNase protection assays revealed that katA andbfrA are on different transcripts, the levels of which are increased by both iron and H₂O₂. Mass spectrometry analysis of whole cells revealed no significant difference in total cellular iron levels in the bfrA,katA, and katA bfrA mutants relative to wild-type bacteria. Our results suggest that P. aeruginosaBfrA may be required as one source of iron for the heme prosthetic group of KatA and thus for protection against H₂O₂.