The Escherichia coli heat shock protease HtrA participates in defense against oxidative stress

The serine protease Htra (DegP), which is indispensable for cell survival a t elevated temperatures, is a peripheral membrane protein, localized on the periplasmic side of the inner membrane in Escherichia coli, and the bioche mical and genetic evidence indicates that the physiological role of HtrA is to degrade denatured proteins formed in the cellular envelope during heat shock. The aim of this study was to find out if the HtrA protease contribut es to protection of the cell against oxidative stress. We compared the infl uence of various oxidizing agents on htrA mutant cells with their effects o n wild-type bacteria, and found that the htr A mutation did not increase se nsitivity to hydrogen peroxide or paraquat but made the cell extremely sens itive to ferrous [Fe(II)] ions, which are known to enhance oxidation of pro teins. Treatment with ferrous ions caused a larger increase in the level of protein carbonyl groups in the membrane fraction of the cell than in the p eriplasm and cytoplasm. Iron-induced oxidation of membrane proteins was enh anced in the htrA mutant relative to wild-type cells. Inhibition of the gro wth of the htrA mutant by iron could be alleviated more efficiently by a ni troxide antioxidant that localizes in the membranes (A-TEMPO) than by a der ivative (40H-TEM-PO) that acts mainly in the soluble fraction of the cell. Inhibition of the growth of the htrA mutant was more pronounced following t reatment with cumene hydroperoxide, which partitions into membranes, than w ith t-butyl hydroperoxide, which forms radical mainly in the cytosol. Both ferrous ions and cumene hydroperoxide, but not hydrogen peroxide, paraquat or t-butyl hydroperoxide, induced synthesis of HtrA. Our results show that HtrA plays a role in defense against oxidative shock and support the hypoth esis that HtrA participates in the degradation of oxidatively damaged prote ins localized in the cell envelope, especially those associated with the me mbranes.