Glc) and catabolite repression mediated by the global regulator cyclic AMP (cAMP)-cAMP receptor protein (CRP). We measured in a systematic way the relation between cellular growth rates and the key parameters of catabolite repression, i.e., the phosphorylated EIIA Crr (EIIA Crr ϳP) level and the cAMP level, using in vitro and in vivo assays. Different growth rates were obtained by using either various carbon sources or by growing the cells with limited concentrations of glucose, sucrose, and mannitol in continuous bioreactor experiments. The ratio of EIIA Crr to EIIA Crr ϳP and the intracellular cAMP concentrations, deduced from the activity of a cAMP-CRP-dependent promoter, correlated well with specific growth rates between 0.3 h ؊1 and 0.7 h ؊1 , corresponding to generation times of about 138 and 60 min, respectively. Below and above this range, these parameters were increasingly uncoupled from the growth rate, which perhaps indicates an increasing role executed by other global control systems, in particular the stringent-relaxed response system.In Escherichia coli, the phosphoenolpyruvate (PEP)-dependent phosphotransferase systems (PTSs) represent important uptake systems for a number of carbohydrates which mediate transport and concomitant phosphorylation of their respective substrates (10,44). In addition to their transport function, all components of the various PTSs of a cell form an important signal transduction system. The signal transduction properties of the PTS depend on the phosphorylation state of its proteins (26,49). The PTSs usually consist of two general proteins, i.e., the PEP-dependent protein kinase enzyme I (EI), and the histidine-containing protein (HPr), and up to 20 different, substrate-specific enzymes II (EII). EII usually comprise two soluble domains EIIA and EIIB involved in phosphotransfer and the membrane-bound transporter domain EIIC (44). The major regulatory output signal of the PTS depends on the phosphorylation level of EIIA Crr (according to its genetic nomenclature), also designated EIIA Glc due to its function as the EIIA domain for the glucose-specific PTS (9, 23, 52). EIIA Crr inhibits the activity of a number of non-PTS transporters and enzymes (8,32,33,35,36), a process referred to as inducer exclusion. Furthermore, the phosphorylated form of EIIA Crr (EIIA Crr ϳP) activates adenylate cyclase (1, 13, 41, 57), which in turn synthesizes cyclic AMP (cAMP) (59). The indicator molecule or alarmone cAMP is the coactivator of the important global transcription factor CRP (cAMP receptor protein). Together, they regulate in a process called cAMP-CRP-dependent catabolite repression efficient transcription of different genes involved in the synthesis of a large number of catabolic enzymes (4,39,43). The central role of EIIA Crr ϳP in the activation of adenylate cyclase is largely based on mutant analysis (13,23,33).The phosphorylation state of the PTS and hence the intracellular cAMP concentrations are postulated to depend largely on two major factors: (i) the uptake rate of any P...
A dynamic mathematical model was developed to describe the uptake of various carbohydrates (glucose, lactose, glycerol, sucrose, and galactose) in Escherichia coli. For validation a number of isogenic strains with defined mutations were used. By considering metabolic reactions as well as signal transduction processes influencing the relevant pathways, we were able to describe quantitatively the phenomenon of catabolite repression in E. coli. We verified model predictions by measuring time courses of several extraand intracellular components such as glycolytic intermediates, EII-A Crr phosphorylation level, both LacZ and PtsG concentrations, and total cAMP concentrations under various growth conditions. The entire data base consists of 18 experiments performed with nine different strains. The model describes the expression of 17 key enzymes, 38 enzymatic reactions, and the dynamic behavior of more than 50 metabolites. The different phenomena affecting the phosphorylation level of EIIA Crr , the key regulation molecule for inducer exclusion and catabolite repression in enteric bacteria, can now be explained quantitatively.Catabolite repression in Escherichia coli designates the observation that if different carbohydrates are present in a medium under unlimited conditions, one of them is usually taken up preferentially. Although the fundamental biochemical principles of the regulatory network have been revealed, a quantitative description of this growth behavior is still missing. The center of the regulatory network is formed by the phosphoenolpyruvate (PEP) 4 :carbohydrate phosphotransferase systems (PTS). These systems are involved in both transport and phosphorylation of a large number of carbohydrates, in movement toward these carbon sources (chemotaxis), and in regulation of a number of metabolic pathways (1-3). The PTS in E. coli consist of two common cytoplasmatic proteins, EI (enzyme I) and HPr (histidine-containing protein), as well as an array of carbohydrate-specific EII (enzyme II) complexes. Because all components of the PTS, depending on their phosphorylation status, can interact with various key regulator proteins, the output of the PTS is represented by the degree of phosphorylation of the proteins involved in phosphoryl group transfer, e.g. unphosphorylated EIIA Crr inhibits the uptake of other non-PTS carbohydrates by a process called inducer exclusion. Phosphorylated EIIA Crr activates the adenylate cyclase (CyaA) and leads to an increase in the intracellular cAMP level.Understanding the regulation of carbohydrate uptake requires a quantitative description of the PTS. In this context it is important that the degree of phosphorylation of EIIA Crr is proportional to the PEP/ pyruvate ratio, when no carbohydrates are transported (4) and the respective equilibrium constant is an upper boundary when the PTS is active (5). The PTS should therefore not be regarded as a measure for the transport of PTS substrates but more as a general measure for carbohydrate availability. One feature of our contribution...
Chemotactic responses in Escherichia coli are typically mediated by transmembrane receptors that monitor chemoeffector levels with periplasmic binding domains and communicate with the flagellar motors through two cytoplasmic proteins, CheA and CheY. CheA autophosphorylates and then donates its phosphate to CheY, which in turn controls flagellar rotation. E. coli also exhibits chemotactic responses to substrates that are transported by the phosphoenolpyruvate (PEP)-dependent carbohydrate phosphotransferase system (PTS). Unlike conventional chemoreception, PTS substrates are sensed during their uptake and concomitant phosphorylation by the cell. The phosphoryl groups are transferred from PEP to the carbohydrates through two common intermediates, enzyme I (El) and phosphohistidine carrier protein (HPr), and then to sugar-specific enzymes II. We found that in mutant strains HPr-like (4) and somehow sensed as chemoeffectors during the uptake process (5, 6). PTSs consist of membraneassociated substrate-specific enzymes II (EIIs) and a common cytoplasmic phosphodonor relay (Fig. 1). EIls are phosphorylated at the expense of PEP through enzyme I (El), a histidine kinase, and a phosphohistidine carrier protein (HPr). During transport of PTS carbohydrates, phosphate groups are transferred through El and HPr to the appropriate ElI and finally to the substrate molecule as it enters the cell (for review, see ref. 7). This phospho-relay activity generates a signal that suppresses clockwise flagellar rotation, thereby extending swimming runs that carry the organism toward higher substrate concentrations (8).The signaling connection between the PTS and MCP chemotactic pathways has long been a mystery. MCPs are not required for PTS chemotaxis, but CheA and CheY are required (9-11), suggesting that PTS signals elicit flagellar responses by modulating phospho-CheY levels, possibly through control of CheA activity (12). E. coli has at least 15 Ells, each of which serves as the "chemoreceptor" for its transport substrates (7). However, neither the binding of substrate to an ElI nor the generation of intracellular carbohydrate-phosphate nor its subsequent degradation is sufficient to trigger a chemotactic response (5,6,10,13,14). In contrast, the common phospho-relay components El and HPr are necessary for uptake of all PTS carbohydrates and for chemotactic responses to them. Conceivably, the flagellar signal derives from an uptake-driven change in phosphate flux through these shared PTS components (6,15). This article describes in vivo and in vitro studies that indicate that the unphosphorylated form of El may be the long-sought missing link between the PTS and MCP phospho-relay circuits. Its signaling target appears to be the CheA kinase.
Glucose is the classical carbon source that is used to investigate the transport, metabolism, and regulation of nutrients in bacteria. Many physiological phenomena like nutrient limitation, stress responses, production of antibiotics, and differentiation are inextricably linked to nutrition. Over the years glucose transport systems have been characterized at the molecular level in more than 20 bacterial species. This review aims to provide an overview of glucose uptake systems found in the eubacterial kingdom. In addition, it will highlight the diverse and sophisticated regulatory features of glucose transport systems.
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