Protoporphyrin (PPIX) efflux by the MacAB-TolC pump in Escherichia coli.
Bottom Line: Escherichia coli regulates heme biosynthesis by a feedback loop involving heme-induced proteolytic cleavage of HemA, glutamyl-tRNA reductase, which is the first enzyme in the heme biosynthetic pathway.We also show that it is disrupted by iron chelation, which reduces the intracellular iron concentration necessary for loading iron into protoporphyrin IX (PPIX, the immediate heme precursor).We propose that PPIX is an endogenous substrate of the MacAB-TolC pump in E. coli and S. typhimurium and that this compound is produced inside bacteria when natural heme homeostasis is disrupted by iron shortage, as happens when bacteria invade the mammalian host.
Affiliation: Unité des Membranes Bactériennes, Département de Microbiologie, Institut Pasteur, 75724, Paris Cedex 15, France.Show MeSH
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Mentions: In the absence of Dip, cultures were non-fluorescent under near-UV light. In the presence of the iron chelator, the entF strain was slightly fluorescent whereas, entF tolC and entF macAB showed strong fluorescence (Fig. 2A). The total porphyrin intracellular contents were determined by fluorescence spectrometry with a near-UV excitation (Fig. 2C). The total intracellular porphyrins in the entF increased 10 times in the presence of iron chelators as compared to the Dip free medium. This tendency was even more pronounced for the entF macAB and entF tolC strains where a boost of 20 and 25, respectively, was achieved in the absence of iron. HPLC analysis of the porphyrin content in the three iron-chelated cultures confirmed these results and showed that the main compound in the cell extract corresponds to PPIX (Fig. 2B). Therefore, in the absence of intracellular iron to ligate PPIX, there was an accumulation of porphyrins including PPIX which is expelled by the MacAB-TolC pump.
Affiliation: Unité des Membranes Bactériennes, Département de Microbiologie, Institut Pasteur, 75724, Paris Cedex 15, France.