PTS 50: Past, Present and Future, or Diauxie Revisited.

Unlabelled: PAST: The title 'PTS 50 or The PTS after 50 years' relies on the first description in 1964 of the phosphoenolpyruvate-dependent carbohydrate:phosphotransferase system (PTS) by Kundig, Gosh and Roseman [Proc Natl Acad Sci USA 1964;52:1067-1074]. The system comprised proteins named Enzyme I, HPr and Enzymes II, as part of a novel PTS for carbohydrates in Gram-negative and Gram-positive bacteria, whose 'biological significance remained unclear'. In contrast, studies which would eventually lead to the discovery of the central role of the PTS in bacterial metabolism had been published since before 1942.

Characterization of pyruvate uptake in Escherichia coli K-12.

The monocarboxylate pyruvate is an important metabolite and can serve as sole carbon source for Escherichia coli. Although specific pyruvate transporters have been identified in two bacterial species, pyruvate transport is not well understood in E. coli.

Bacterial PEP-dependent carbohydrate: phosphotransferase systems couple sensing and global control mechanisms.

The PEP-dependent carbohydrate:phosphotransferase systems (PTSs) of enteric bacteria constitute a complex sensory system which involves as its central element a PEP-dependent His-protein kinase (Enzyme I). As a unit, the PTS comprises up to 20 different transporters per cell which correspond to its chemoreceptors for PTS carbohydrates, and several targeting subunits, which include in the low [G+C] Gram-positive bacteria an ancillary Ser/Thr-protein kinase. The PTS senses the presence of carbohydrates, in particular glucose, in the medium and the energy state of the cell, in the form of either the intracellular PEP-to-pyruvate ratio or the D-fructose-bisphosphate levels.

Correlation between growth rates, EIIACrr phosphorylation, and intracellular cyclic AMP levels in Escherichia coli K-12.

In Escherichia coli K-12, components of the phosphoenolpyruvate-dependent phosphotransferase systems (PTSs) represent a signal transduction system involved in the global control of carbon catabolism through inducer exclusion mediated by phosphoenolpyruvate-dependent protein kinase enzyme IIA(Crr) (EIIA(Crr)) (= EIIA(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 genes and enzymes for the catabolism of galactitol, D-tagatose, and related carbohydrates in Klebsiella oxytoca M5a1 and other enteric bacteria display convergent evolution.

Enteric bacteria (Enteriobacteriaceae) carry on their single chromosome about 4000 genes that all strains have in common (referred to here as "obligatory genes"), and up to 1300 "facultative" genes that vary from strain to strain and from species to species. In closely related species, obligatory and facultative genes are orthologous genes that are found at similar loci. We have analyzed a set of facultative genes involved in the degradation of the carbohydrates galactitol, D-tagatose, D-galactosamine and N-acetyl-galactosamine in various pathogenic and non-pathogenic strains of these bacteria.

Mutations which uncouple transport and phosphorylation in the D-mannitol phosphotransferase system of Escherichia coli K-12 and Klebsiella pneumoniae 1033-5P14.

Mutants of Escherichia coli K-12 were isolated which lack the normal phosphotransferase system-dependent catabolic pathway for D-mannitol (Mtl). In some mutants the pts genes for the general proteins enzyme I and histidine protein of the phosphoenolpyruvate-dependent carbohydrate phosphotransferase systems were deleted. Other mutants expressed truncated mannitol-specific enzymes II (II(Mtl)) which lacked the IIA(Mtl) or IIBA(Mtl) domain(s), and the mtlA genes originated either from E.

Adaptation of sucrose metabolism in the Escherichia coli wild-type strain EC3132.

Although Escherichia coli strain EC3132 possesses a chromosomally encoded sucrose metabolic pathway, its growth on low sucrose concentrations (5 mM) is unusually slow, with a doubling time of 20 h. In this report we describe the subcloning and further characterization of the corresponding csc genes and adjacent genes. The csc regulon comprises three genes for a sucrose permease, a fructokinase, and a sucrose hydrolase (genes cscB, cscK, and cscA, respectively).

Two class II D-tagatose-bisphosphate aldolases from enteric bacteria.

Escherichia coli, Salmonella enterica, Klebsiella pneumoniaeand Klebsiella oxytocawere found to contain two D-tagatose 1,6-bisphosphate (TagBP)-specific aldolases involved in catabolism of galactitol (genes gatY gatZ) and of N-acetyl-galactosamine and D-galactosamine (genes kbaY kbaZ,also called agaY agaZ). The two aldolases were closely related (> or = 53.8% identical amino acids) and could substitute for each other in vivo.

The organization of metabolic reaction networks. III. Application for diauxic growth on glucose and lactose.

A mathematical model to describe carbon catabolite repression in Escherichia coli is developed and in part validated. The model is aggregated from two functional units describing glucose and lactose transport and degradation. Both units are members of the crp modulon and are under control of a global signal transduction system which calculates the signals that turn on or off gene expression for the specific enzymes.

Modeling of inducer exclusion and catabolite repression based on a PTS-dependent sucrose and non-PTS-dependent glycerol transport systems in Escherichia coli K-12 and its experimental verification.

We used genetically engineered sucrose positive Escherichia coli K-12 derivatives as a model system for the modeling and experimental verification of regulatory processes in bacteria. These cells take up and metabolize sucrose by the phosphoenolpyruvate (PEP)-dependent sucrose phosphotransferase system (Scr-PTS). Expression of the scr genes, which cluster in two different operons (scrYAB and scrK), is negatively controlled by the ScrR repressor.

The mtl genes and the mannitol-1-phosphate dehydrogenase from Klebsiella pneumoniae KAY2026.

The mtl operon of Klebsiella pneumoniae KAY2026 (formerly Aerobacter aerogenes 1033-5P14) was shown to contain as the promoter-proximal gene mtlA, encoding a D-mannitol-specific enzyme II transporter (IICBA(Mtl)). This gene is followed by mtlD, coding for a mannitol-1-phosphate dehydrogenase (MtlD, 382 amino acid residues), and mtlR (MtlR, 195 amino acid residues) coding for a putative repressor, gene mtlR overlaps the termination codon of mtlD. The DNA and protein sequences are highly similar to the corresponding genes (81% identical bp) and proteins (79-85% identical amino acids) of Escherichia coli K-12.

Metabolic networks: a signal-oriented approach to cellular models.

Complete genomes, far advanced proteomes, and even 'metabolomes' are available for at least a few organisms, e.g., Escherichia coli.

The organization of metabolic reaction networks: a signal-oriented approach to cellular models.

Complex metabolic networks are characterized by a great number of elements and many regulatory loops. The description of these networks with mathematical models requires the definition of functional units that group together several cellular processes. The approach presented here is based on the idea that cellular functional units may be assigned directly to mathematical modeling objects.

Pathways for the utilization of N-acetyl-galactosamine and galactosamine in Escherichia coli.

Among enteric bacteria, the ability to grow on N-acetyl-galactosamine (GalNAc or Aga) and on D-galactosamine (GalN or Gam) differs. Thus, strains B, C and EC3132 of Escherichia coli are Aga+ Gam+ whereas E. coli K-12 is Aga- Gam-, similarly to Klebsiella pneumoniae KAY2026, Klebsiella oxytoca M5a1 and Salmonella typhimurium LT2.

Glucose transporter mutants of Escherichia coli K-12 with changes in substrate recognition of IICB(Glc) and induction behavior of the ptsG gene.

In Escherichia coli K-12, the major glucose transporter with a central role in carbon catabolite repression and in inducer exclusion is the phosphoenolpyruvate-dependent glucose:phosphotransferase system (PTS). Its membrane-bound subunit, IICB(Glc), is encoded by the gene ptsG; its soluble domain, IIA(Glc), is encoded by crr, which is a member of the pts operon. The system is inducible by D-glucose and, to a lesser degree, by L-sorbose.

Elucidation of a PTS-carbohydrate chemotactic signal pathway in Escherichia coli using a time-resolved behavioral assay.

Chemotaxis of Escherichia coli toward phosphotransferase systems (PTSs)-carbohydrates requires phosphoenolpyruvate-dependent PTSs as well as the chemotaxis response regulator CheY and its kinase, CheA. Responses initiated by flash photorelease of a PTS substrates D-glucose and its nonmetabolizable analog methyl alpha-D-glucopyranoside were measured with 33-ms time resolution using computer-assisted motion analysis. This, together with chemotactic mutants, has allowed us to map out and characterize the PTS chemotactic signal pathway.

Site-directed mutagenesis of loop L3 of sucrose porin ScrY leads to changes in substrate selectivity.

The difference in substrate selectivity of the maltodextrin (LamB) and sucrose (ScrY) porins is attributed mainly to differences in loop L3, which is supposed to constrict the lumen of the pores. We show that even a single mutation (D201Y) in loop L3 leads to a narrowing of the substrate range of ScrY to that resembling LamB. In addition, we removed the putative N-terminal coiled-coil structure of ScrY and studied the effect of this deletion on sucrose transport.

Genes for D-arabinitol and ribitol catabolism from Klebsiella pneumoniae.

The enzymes for catabolism of the pentitols D-arabinitol (Dal) and ribitol (Rbt) and the corresponding genes from Klebsiella pneumoniae (dal and rbt) and Escherichia coli (atl and rtl) have been used intensively in experimental evolutionary studies. Four dal and four rbt genes from the chromosome of K. pneumoniae 1033-5P14 were cloned and sequenced.

Substrate recognition domains as revealed by active hybrids between the D-arabinitol and ribitol transporters from Klebsiella pneumoniae.

Two new genes, dalT and rbtT, have been cloned from the dal operon for D-arabinitol and the rbt operon for ribitol uptake and degradation, respectively, in Klebsiella pneumoniae 1033-5P14, derivative KAY2026. Each gene codes for a specific transporter which, based on sequence data, belongs to a large family of carbohydrate transporters which constitutes 12 transmembrane helices. DalT and RbtT show an unusually high similarity (86.

Identification of a new porin, RafY, encoded by raffinose plasmid pRSD2 of Escherichia coli.

The conjugative plasmid pRSD2 carries a raf operon that encodes a peripheral raffinose metabolic pathway in enterobacteria. In addition to the previously known raf genes, we identified another gene, rafY, which in Escherichia coli codes for an outer membrane protein (molecular mass, 53 kDa) similar in function to the known glycoporins LamB (maltoporin) and ScrY (sucrose porin). Sequence comparisons with LamB and ScrY revealed no significant similarities; however, both lamB and scrY mutants are functionally complemented by RafY.

CTnscr94, a conjugative transposon found in enterobacteria.

Conjugational transposons are important for horizontal gene transfer in gram-positive and gram-negative bacteria, but have not been reported yet for enteric bacteria. Salmonella senftenberg 5494-57 has previously been shown to transfer by conjugation genes for a sucrose fermentation pathway which were located on a DNA element called scr-94. We report here that the corresponding scr genes for a phosphoenolpyruvate-dependent sucrose:phosphotransferase system and a sucrose metabolic pathway are located on a large (ca.

Molecular analysis of the gat genes from Escherichia coli and of their roles in galactitol transport and metabolism.

In enteric bacteria, the hexitol galactitol (Gat) (formerly dulcitol) is taken up through enzyme II (II(Gat)) of the phosphoenolpyruvate-dependent phosphotransferase system (PTS), and accumulated as galactitol 1-phosphate (Gat1P). The gat genes involved in galactitol metabolism have been isolated from the wild-type isolate Escherichia coli EC3132 and cloned on a 7.8-kbp PstI DNA fragment.

Molecular analysis of the scrA and scrB genes from Klebsiella pneumoniae and plasmid pUR400, which encode the sucrose transport protein Enzyme II Scr of the phosphotransferase system and a sucrose-6-phosphate invertase.

The Klebsiella pneumoniae genes scrA and scrB are indispensable for sucrose (Scr) utilisation. Gene scrA codes for an Enzyme IIScr (IIScr) transport protein of the phosphoenolpyruvate-dependent carbohydrate: phosphotransferase system (PTS), while scrB encodes a sucrose 6-phosphate specific invertase. A 3.

Coupling the phosphotransferase system and the methyl-accepting chemotaxis protein-dependent chemotaxis signaling pathways of Escherichia coli.

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.