Physiological and Proteomic Analysis of Escherichia coli Iron-Limited Chemostat Growth

Abstract

Iron bioavailability is a major limiter of bacterial growth in mammalian host tissue and thus represents an important area of study. Escherichia coli K-12 metabolism was studied at four levels of iron limitation in chemostats using physiological and proteomic analyses. The data documented an E. coli acclimation gradient where progressively more severe iron scarcity resulted in a larger percentage of substrate carbon being directed into an overflow metabolism accompanied by a decrease in biomass yield on glucose. Acetate was the primary secreted organic by-product for moderate levels of iron limitation, but as stress increased, the metabolism shifted to secrete primarily lactate (ϳ70% of catabolized glucose carbon). Proteomic analysis reinforced the physiological data and quantified relative increases in glycolysis enzyme abundance and decreases in tricarboxylic acid (TCA) cycle enzyme abundance with increasing iron limitation stress. The combined data indicated that E. coli responds to limiting iron by investing the scarce resource in essential enzymes, at the cost of catabolic efficiency (i.e., downregulating high-ATP-yielding pathways containing enzymes with large iron requirements, like the TCA cycle). Acclimation to iron-limited growth was contrasted experimentally with acclimation to glucose-limited growth to identify both general and nutrient-specific acclimation strategies. While the iron-limited cultures maximized biomass yields on iron and increased expression of iron acquisition strategies, the glucose-limited cultures maximized biomass yields on glucose and increased expression of carbon acquisition strategies. This study quantified ecologically competitive acclimations to nutrient limitations, yielding knowledge essential for understanding medically relevant bacterial responses to host and to developing intervention strategies.

Physiological, biomass elemental composition and proteomic analyses of Escherichia coli ammonium-limited chemostat growth, and comparison with iron- and glucose-limited chemostat growth

“…Individual aliquots of E. coli K-12 MG1655 were stored frozen at 280 uC in 25% glycerol. Media details were described previously (Folsom et al, 2014 A summary of the medium variations used to achieve ammonium-, iron-or glucose-limited conditions can be found in Table 1. Theoretical medium design calculations can be found in File S1 (available in the online Supplementary Material).…”

“…Chemostats were operated as described previously (Folsom et al, 2014). Briefly, chemostats with 300 ml of medium were inoculated with 450 ml of E. coli K-12 MG1655 freezer stock and incubated without medium flow for at least 6 h at 37 uC until the reactors were turbid.…”

“…Here, we report E. coli physiology, biomass elemental content and central metabolism proteome for ammoniumlimited chemostat growth at four dilution rates, and contrast the data with published iron-and glucose-limited growth at the same dilution rates using the same E. coli K-12 strain, M9 medium and reactor configuration (Folsom et al, 2014). The previously reported data were leveraged to gain additional physiological knowledge by noting nutrient-specific and general stress responses.…”

“…Iron limitation is another important bacterial nutrient-limitation stress. It is central in medical infections and to marine microbial growth, and has been described recently by our group (Folsom et al, 2014).…”

Vibrio Iron Transport: Evolutionary Adaptation to Life in Multiple Environments

“…While glycolysis does not require iron-containing enzymes, other pathways, such as the Entner-Doudoroff pathway and the TCA cycle, have enzymes that require iron for activity. Under low-iron conditions, E. coli shifts its metabolism to increase glycolysis and decrease the level of TCA cycle enzymes (244). It is likely that vibrios alter their metabolism in a similar way.…”