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Abstract Metabolic cross-feeding plays vital roles in promoting ecological diversity. While some microbes depend on exchanges of essential nutrients for growth, the forces driving the extensive cross-feeding needed to support the coexistence of free-living microbes are poorly understood. Here we characterize bacterial physiology under self-acidification and establish that extensive excretion of key metabolites following growth arrest provides a collaborative, inter-species mechanism of stress resistance. This collaboration occurs not only between species isolated from the same community, but also between unrelated species with complementary (glycolytic vs. gluconeogenic) modes of metabolism. Cultures of such communities progress through distinct phases of growth-dilution cycles, comprising of exponential growth, acidification-triggered growth arrest, collaborative deacidification, and growth recovery, with each phase involving different combinations of physiological states of individual species. Our findings challenge the steady-state view of ecosystems commonly portrayed in ecological models, offering an alternative dynamical view based on growth advantages of complementary species in different phases. 

Abstract Protein serine/threonine/tyrosine (S/T/Y) phosphorylation is an essential and frequent post-translational modification in eukaryotes, but historically has been considered less prevalent in bacteria because fewer proteins were found to be phosphorylated and most proteins were modified to a lower degree. Recent proteomics studies greatly expanded the phosphoproteome of Escherichia coli to more than 2000 phosphorylation sites (phosphosites), yet mechanisms of action were proposed for only six phosphosites and fitness effects were described for 38 phosphosites upon perturbation. By systematically characterizing functional relevance of S/T/Y phosphorylation in E. coli metabolism, we found 44 of the 52 mutated phosphosites to be functional based on growth phenotypes and intracellular metabolome profiles. By effectively doubling the number of known functional phosphosites, we provide evidence that protein phosphorylation is a major regulation process in bacterial metabolism. Combining in vitro and in vivo experiments, we demonstrate how single phosphosites modulate enzymatic activity and regulate metabolic fluxes in glycolysis, methylglyoxal bypass, acetate metabolism and the split between pentose phosphate and Entner-Doudoroff pathways through mechanisms that include shielding the substrate binding site, limiting structural dynamics, and disrupting interactions relevant for activity in vivo.