Metal cofactors in regulatory proteins and enzymes can serve as sensors for oxygen and oxidative stress. Reactive oxygen species interact with the bound metal ions and cause oxidative damage to amino acids surrounding the ligand-binding site. We are studying how oxidative damage can target specific proteins to be efficiently degraded by the AAA+ proteases ClpXP, ClpAP and Lon. Damage-specific substrates under investigation include two environmentally sensitive transcription regulators, PerR and FNR, and ALA synthase, a critical enzyme in heme biosynthesis and red blood cell development. A well-characterized example of damage-triggered degradation is control of the ubiquitous bacterial hydrogen peroxide sensor, PerR. Oxidation of PerR at its regulatory metal-binding site serves as the peroxide sensor and irreversibly modifies two histidines such that they are unable to be repaired. PerR is thus a “suicide regulator” that has to be degraded and resynthesized for each round of transcriptional regulation. We find that Lon preferentially degrades oxidized PerR both <i>in vivo</i> and <i>in vitro</i>. By mapping a specific peptide sequence required for Lon recognition of PerR we found that this Lon-binding region is exposed to solvent in oxidized PerR, but is sustainably buried in the metal-bound, undamaged form. These insights provide a compelling mechanism explaining how the metal-dependent, oxidation induced conformational change in PerR is critical for protease resistance verses sensitivity; similar mechanisms may control damaged-induced proteolysis of other critical proteins.

How metal-dependent oxidation can redirect AAA+ proteases to new protein substrates

253rd National Meeting (2017)

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