Metabolism of long-chain fatty acids affects disulfide bond formation in Escherichia coli and activates envelope stress response pathways as a combat strategy

The envelope of gram-negative bacteria serves as the first line of defense against environmental insults. Therefore, its integrity is continuously monitored and maintained by several envelope stress response (ESR) systems. Due to its oxidizing environment, the envelope represents an important site for disulfide bond formation. In Escherichia coli, the periplasmic oxidoreductase, DsbA introduces disulfide bonds in substrate proteins and transfers electrons to the inner membrane oxidoreductase, DsbB. Under aerobic conditions, the reduced form of DsbB is re-oxidized by ubiquinone, an electron carrier in the electron transport chain (ETC). Given the critical role of ubiquinone in transferring electrons derived from the oxidation of reduced cofactors, we were intrigued whether metabolic conditions that generate a large number of reduced cofactors render ubiquinone unavailable for disulfide bond formation. To test this, here we investigated the influence of metabolism of long-chain fatty acid (LCFA), an energy-rich carbon source, on the redox state of the envelope. We show that LCFA degradation increases electron flow in the ETC. Further, we find that whereas cells metabolizing LCFAs exhibit several characteristics of insufficient disulfide bond formation, these hallmarks are averted in cells exogenously provided with ubiquinone. Importantly, the ESR pathways, Cpx and σE, are activated by envelope signals generated during LCFA metabolism, and these systems maintain proper disulfide bond formation. We find that σE downregulation hampers disulfide bond formation only in the absence of Cpx, and amongst the two ESR systems, only Cpx senses redox-dependent signal and is induced to a greater extent by LCFAs. Therefore, we argue that Cpx is the primary ESR that senses and maintains envelope redox homeostasis. Taken together, our results demonstrate an intricate relationship between cellular metabolism and disulfide bond formation dictated by ETC and ESR, and provide the basis for examining whether similar mechanisms control envelope redox status in other gram-negative bacteria.Author summaryDisulfide bonds contribute to the folding and stability of many extracytoplasmic proteins in all domains of life. In gram-negative bacteria, including Escherichia coli, disulfide bond formation occurs in the oxidizing environment of the periplasmic space enclosed within the outer and inner membrane layers of the envelope. Because disulfide-bonded proteins are involved in diverse biological processes, bacteria must monitor the envelope redox status and elicit an appropriate response when perturbations occur; however, these mechanisms are not well elucidated. Here, we demonstrated that the metabolism of an energy-rich carbon source, long-chain fatty acid (LCFA) hampers disulfide bond formation in E. coli. An envelope stress response (ESR) system, Cpx, senses this redox imbalance and maintains proper disulfide bond formation. The σE pathway, another ESR system, plays an ancillary role in maintaining redox homeostasis. LCFA metabolism, disulfide bond formation, and ESR systems have independently been implicated in the pathogenesis of several gram-negative bacteria. The present study sets the basis to explore whether LCFA metabolism impacts the virulence of these bacteria by influencing the redox status of their envelope and activation of ESR pathways.