CTCF/cohesin organize the ground state of chromatin-nuclear speckle association.

The interchromatin space in the cell nucleus contains various membrane-less nuclear bodies. Recent findings indicate that nuclear speckles, comprising a distinct nuclear body, exhibit interactions with certain chromatin regions in a ground state. Key questions are how this ground state of chromatin-nuclear speckle association is established and what are the gene regulatory roles of this layer of nuclear organization.

Polycomb-lamina antagonism partitions heterochromatin at the nuclear periphery.

The genome can be divided into two spatially segregated compartments, A and B, which partition active and inactive chromatin states. While constitutive heterochromatin is predominantly located within the B compartment near the nuclear lamina, facultative heterochromatin marked by H3K27me3 spans both compartments. How epigenetic modifications, compartmentalization, and lamina association collectively maintain heterochromatin architecture remains unclear.

Implementation of the INTERGROWTH-21st Project in the UK.

There are approximately 10,000 births per year in the county of Oxfordshire in the UK, which is one of the two European sites for the International Fetal and Newborn Growth Consortium for the 21(st) Century (INTERGROWTH-21(st) ) Project. The samples for both components of the project--the Fetal Growth Longitudinal Study (FGLS) and Newborn Cross-Sectional Study (NCSS)--were drawn from the John Radcliffe Hospital, a major university hospital with a large regional role that covers more than 75% of deliveries in the county. Special activities to encourage participation in this population included the formation of a research coalition to streamline recruitment in the Maternity Unit and the distribution of study information leaflets to women using the hospital's antenatal care service.

The chitin catabolic cascade in the marine bacterium Vibrio cholerae: characterization of a unique chitin oligosaccharide deacetylase.

Chitin, one of the most abundant organic substances in nature, is consumed by marine bacteria, such as Vibrio cholerae, via a multitude of tightly regulated genes (Li and Roseman 2004, Proc Natl Acad Sci USA. 101:627-631). One such gene, cod, is reported here.

The monomer/dimer transition of enzyme I of the Escherichia coli phosphotransferase system.

Enzyme I (EI) is the first protein in the phosphotransfer sequence of the bacterial phosphoenolpyruvate:glycose phosphotransferase system. This system catalyzes sugar phosphorylation/transport and is stringently regulated. Since EI homodimer accepts the phosphoryl group from phosphoenolpyruvate (PEP), whereas the monomer does not, EI may be a major factor in controlling sugar uptake.

Properties of the C-terminal domain of enzyme I of the Escherichia coli phosphotransferase system.

The bacterial phosphoenolpyruvate (PEP):glycose phosphotransferase system (PTS) mediates uptake/phosphorylation of sugars. The transport of all PTS sugars requires Enzyme I (EI) and a phosphocarrier histidine protein of the PTS (HPr). The PTS is stringently regulated, and a potential mechanism is the monomer/dimer transition of EI, because only the dimer accepts the phosphoryl group from PEP.

Effects of mutations and truncations on the kinetic behavior of IIAGlc, a phosphocarrier and regulatory protein of the phosphoenolpyruvate phosphotransferase system of Escherichia coli.

IIAGlc, a component of the glucose-specific phosphoenolpyruvate:phosphotransferase system (PTS) of Escherichia coli, is important in regulating carbohydrate metabolism. In Glc uptake, the phosphotransfer sequence is: phosphoenolpyruvate --> Enzyme I --> HPr --> IIAGlc --> IICBGlc --> Glc. (HPr is the first phosphocarrier protein of the PTS.

Ultrafast fluorescence dynamics of tryptophan in the proteins monellin and IIAGlc.

The complete time-resolved fluorescence of tryptophan in the proteins monellin and IIA(Glc) has been investigated, using both an upconversion spectrophotofluorometer with 150 fs time resolution and a time-correlated single photon counting apparatus on the 100 ps to 20 ns time scale. In monellin, the fluorescence decay displays multiexponential character with decay times of 1.2 and 16 ps, and 0.

Transient state kinetics of enzyme IICBGlc, a glucose transporter of the phosphoenolpyruvate phosphotransferase system of Escherichia coli: equilibrium and second order rate constants for the glucose binding and phosphotransfer reactions.

During translocation across the cytoplasmic membrane of Escherichia coli, glucose is phosphorylated by phospho-IIA(Glc) and Enzyme IICB(Glc), the last two proteins in the phosphotransfer sequence of the phosphoenolpyruvate:glucose phosphotransferase system. Transient state (rapid quench) methods were used to determine the second order rate constants that describe the phosphotransfer reactions (phospho-IIA(Glc) to IICB(Glc) to Glc) and also the second order rate constants for the transfer from phospho-IIA(Glc) to molecularly cloned IIB(Glc), the soluble, cytoplasmic domain of IICB(Glc). The rate constants for the forward and reverse phosphotransfer reactions between IIA(Glc) and IICB(Glc) were 3.

Transient state kinetics of Enzyme I of the phosphoenolpyruvate:glycose phosphotransferase system of Escherichia coli: equilibrium and second-order rate constants for the phosphotransfer reactions with phosphoenolpyruvate and HPr.

The first two reactions in the phosphotransfer sequence of bacterial phosphoenolpyruvate:glycose phosphotransferase systems are the autophosphorylation of Enzyme I by phosphoenolpyruvate followed by the transfer of the phospho group to the low-molecular weight protein, HPr. Transient state kinetic methods were used to estimate the second-order rate constants for both phosphotransfer reactions. These measurements support previous conclusions that only the dimer of Enzyme I, EI2, is autophosphorylated, and that the rate of formation of dimer is slow compared to the rate of its phosphorylation.

Subcellular distribution of enzyme I of the Escherichia coli phosphoenolpyruvate:glycose phosphotransferase system depends on growth conditions.

The phosphoenolpyruvate:glycose phosphotransferase system (PTS) participates in important functions in the bacterial cell, including the phosphorylation/uptake of PTS sugars. Enzyme I (EI), the first protein of the PTS complex, accepts the phosphoryl group from phosphoenolpyruvate, which is then transferred through a chain of proteins to the sugar. In these studies, a mutant GFP, enhanced yellow fluorescent protein (YFP), was linked to the N terminus of EI, giving Y-EI.

The Vibrio cholerae chitin utilization program.

Chitin, an insoluble polymer of GlcNAc, is an abundant source of carbon, nitrogen, and energy for marine microorganisms. Microarray expression profiling and mutational studies of Vibrio cholerae growing on a natural chitin surface, or with the soluble chitin oligosaccharides (GlcNAc)(2-6), GlcNAc, or the glucosamine dimer (GlcN)2 identified three sets of differentially regulated genes. We show that (i) ChiS, a sensor histidine kinase, regulates expression of the (GlcNAc)(2-6) gene set, including a (GlcNAc)2 catabolic operon, two extracellular chitinases, a chitoporin, and a PilA-containing type IV pilus, designated ChiRP (chitin-regulated pilus) that confers a significant growth advantage to V.

The chitinolytic cascade in Vibrios is regulated by chitin oligosaccharides and a two-component chitin catabolic sensor/kinase.

Chitin, a highly insoluble polymer of GlcNAc, is produced in massive quantities in the marine environment. Fortunately for survival of aquatic ecosystems, chitin is rapidly catabolized by marine bacteria. Here we describe a bacterial two-component hybrid sensor/kinase (of the ArcB type) that rigorously controls expression of approximately 50 genes, many involved in chitin degradation.

Molecular cloning and characterization of a unique beta-glucosidase from Vibrio cholerae.

We have recently reported the molecular cloning of a gene, gspK, in Vibrio cholerae that encodes a specific glucosamine kinase. We describe here the identification of bglA, a gene contiguous to gspK in a presumptive large chitin catabolic operon. BglA was molecularly cloned into Escherichia coli, and the protein BglA was overexpressed and purified to apparent homogeneity.

Isolation of a glucosamine-specific kinase, a unique enzyme of Vibrio cholerae.

We showed previously that chitin catabolism by the marine bacterium Vibrio furnissii involves at least three signal transduction systems and many genes, several of which were molecularly cloned, and the corresponding proteins were characterized. The predicted amino acid sequences of these proteins showed a high degree of identity to the corresponding proteins from Vibrio cholerae, whose complete genomic sequence has recently been determined. We have therefore initiated studies with V.

IIA(Glc) allosteric control of Escherichia coli glycerol kinase: binding site cooperative transitions and cation-promoted association by Zinc(II).

The catalytic activity of glycerol kinase (EC 2.7.1.

Analytical sedimentation of the IIAChb and IIBChb proteins of the Escherichia coli N,N'-diacetylchitobiose phosphotransferase system. Demonstration of a model phosphotransfer transition state complex.

The phosphoenolpyruvate:glycose transferase system (PTS) is a prototypic signaling system responsible for the vectorial uptake and phosphorylation of carbohydrate substrates. The accompanying papers describe the proteins and product of the Escherichia coli N, N-diacetylchitobiose ((GlcNAc)(2)) PTS-mediated permease. Unlike most PTS transporters, the Chb system is composed of two soluble proteins, IIA(Chb) and IIB(Chb), and one transmembrane receptor (IIC(Chb)).

The transport/phosphorylation of N,N'-diacetylchitobiose in Escherichia coli. Characterization of phospho-IIB(Chb) and of a potential transition state analogue in the phosphotransfer reaction between the proteins IIA(Chb) AND IIB(Chb).

Enzyme II permeases of the phosphoenolpyruvate:glycose phosphotransferase system comprise one to five separately encoded polypeptides, but most contain similar domains (IIA, IIB, and IIC). The phosphoryl group is transferred from one domain to another, with histidine as the phosphoryl acceptor in IIA and cysteine as the acceptor in certain IIB domains. IIB(Chb) is a phosphocarrier in the uptake/phosphorylation of the chitin disaccharide, (GlcNAc)(2) by Escherichia coli and is unusual because it is separately encoded and soluble.

Isolation and characterization of IIAChb, a soluble protein of the enzyme II complex required for the transport/phosphorylation of N, N'-diacetylchitobiose in Escherichia coli.

N,N'-Diacetylchitobiose is transported/phosphorylated in Escherichia coli by the (GlcNAc)(2)-specific Enzyme II permease of the phosphoenolpyruvate:glycose phosphotransferase system. IIA(Chb), one protein of the Enzyme II complex, was cloned and purified to homogeneity. IIA(Chb) and phospho-IIA(Chb) form stable homodimers ().

The chitin disaccharide, N,N'-diacetylchitobiose, is catabolized by Escherichia coli and is transported/phosphorylated by the phosphoenolpyruvate:glycose phosphotransferase system.

We have previously reported that wild type strains of Escherichia coli grow on the chitin disaccharide N,N'-diacetylchitobiose, (GlcNAc)(2), as the sole source of carbon (Keyhani, N. O., and Roseman, S.

Chitin catabolism in the marine bacterium Vibrio furnissii. Identification, molecular cloning, and characterization of A N, N'-diacetylchitobiose phosphorylase.

The major product of bacterial chitinases is N,N'-diacetylchitobiose or (GlcNAc)(2). We have previously demonstrated that (GlcNAc)(2) is taken up unchanged by a specific permease in Vibrio furnissii (unlike Escherichia coli). It is generally held that marine Vibrios further metabolize cytoplasmic (GlcNAc)(2) by hydrolyzing it to two GlcNAcs (i.

Chitin catabolism in the marine bacterium Vibrio furnissii. Identification and molecular cloning of a chitoporin.

Chitin catabolism by the marine bacterium Vibrio furnissii involves many genes and proteins, including two unique periplasmic hydrolases, a chitodextrinase and a beta-N-acetylglucosaminidase (Keyhani, N. O. , and Roseman, S.

Understanding glucose transport by the bacterial phosphoenolpyruvate:glycose phosphotransferase system on the basis of kinetic measurements in vitro.

The kinetic parameters in vitro of the components of the phosphoenolpyruvate:glycose phosphotransferase system (PTS) in enteric bacteria were collected. To address the issue of whether the behavior in vivo of the PTS can be understood in terms of these enzyme kinetics, a detailed kinetic model was constructed. Each overall phosphotransfer reaction was separated into two elementary reactions, the first entailing association of the phosphoryl donor and acceptor into a complex and the second entailing dissociation of the complex into dephosphorylated donor and phosphorylated acceptor.