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J Delgado

Publications and source records attributed to J Delgado.

At least 19 recordsLinked to original sources
PubMed1992

Dynamic metabolic control theory. A methodology for investigating metabolic regulation using transient metabolic data.

The purposes of the dynamic metabolic control theory are to provide a theoretical basis for estimating the control coefficients using the transient metabolic responses and to gain insights into the metabolic regulation in the transient states. The numerical application of this theory is relatively straightforward: it involves a standard linear regression and a matrix multiplication. Although the equations are exact only for linear kinetics, they yield relatively good estimates of the control coefficients for nonlinear systems.

Cells↗
PubMed1992

Metabolic control analysis using transient metabolite concentrations. Determination of metabolite concentration control coefficients.

The methodology previously developed for determining the Flux Control Coefficients [Delgado & Liao (1992) Biochem. J. 282, 919-927] is extended to the calculation of metabolite Concentration Control Coefficients. It is shown that the transient metabolite concentrations are related by a few algebraic equations, attributed to mass balance, stoichiometric constraints, quasi-equilibrium or quasi-steady states, and kinetic regulations. The coefficients in these relations can be estimated using linear regression, and can be used to calculate the Control Coefficients. The theoretical basis and two examples are discussed. Although the methodology is derived based on the linear approximation of enzyme kinetics, it yields reasonably good estimates of the Control Coefficients for systems with non-linear kinetics.

Enzymes↗
PubMed1992

Determination of Flux Control Coefficients from transient metabolite concentrations.

Flux Control Coefficients have been used in the analysis of metabolic regulation for quantifying the effect of an enzyme on the overall steady-state flux. However, the experimental determination of these coefficients is very time-consuming, involving either determining the individual enzyme kinetics or perturbing the enzyme activity by genetic or other means. We developed a methodology that enables the determination of the Flux Control Coefficients from transient metabolite concentrations without knowing kinetic parameters. The transient states can be generated by changing the incubation conditions or adding the initial substrate. This approach is suitable for investigating metabolic regulation in vivo or multiple enzyme systems in vitro. It is particularly helpful if used in conjunction with n.m.r. measurements. The approach is based on a relationship between transient metabolite concentrations and the Flux Control Coefficients. The methodology has been improved from our previous results, and it is illustrated by three examples with simple pathway topologies.

Enzymes↗
PubMed1991

Ca2(+)-enhanced phosphorylation of a chimeric protein kinase involved with bacterial signal transduction.

The Tar-EnvZ hybrid molecule (Taz1) is an inner membrane transducer that activates OmpR, a transcriptional activator for porin gene expression (ompC), in response to an aspartic acid signal. Signal transduction by Taz1 most likely involves a phosphorylated Taz1 intermediate that donates its phosphate to OmpR. Phosphorylated OmpR has already been implicated in transcriptional activation of porin genes. Using a cell-free system containing Taz1-enriched membrane fractions, we have examined the phosphorylation properties of Taz1 and the stimulatory effects of divalent and monovalent ions. Highest activation of Taz1 phosphorylation was observed with CaCl2, and its stimulation could be observed with as low as 60 microM of CaCl2. Phosphorylated Taz1 could readily donate its phosphate group to OmpR in the presence of calcium. CaCl2 was also able to enhance phosphorylation of intact membrane-bound EnvZ and a cytoplasmic fragment of EnvZ lacking the receptor and transmembrane domains. These results indicate that the site for CaCl2 stimulation is within the cytoplasmic region of EnvZ and probably involves an enhanced rate of EnvZ phosphorylation.

Autoradiography↗
PubMed1990

Differential diagnosis of Taenia saginata and Taenia solium with DNA probes.

A size selected genomic DNA library was constructed using DNA extracted from Taenia saginata. The DNA was digested using the restriction enzyme EcoR1 under star conditions and the 2-4 kbase fraction, selected following sucrose density-gradient separation, was cloned in the bacteriophage lambda gt 10. A panel of cestode DNAs including Taenia saginata, Taenia solium, Taenia taeniaeformis, Taenia crassiceps, Echinococcus granulosus and DNAs of bovine, porcine and human origin were used in conjunction with hybridization analysis to identify two recombinant bacteriophages. The first probe, designated HDP1, reacted specifically with T. saginata DNA. The second, designated HDP2, reacted with DNA from both T. saginata and T. solium but not the other DNA samples and thus provided a general reagent for positive identification of fragments of Taenia spp. proglottides of human faecal origin. If used in conjunction the two DNA probes allow positive identification of T. saginata. In the clinical situation it is important to be able to distinguish T. saginata and T. solium infections and DNA probes such as these may be useful in such differentiation.

Animals↗
PubMed1990

In vivo phosphorylation of OmpR, the transcription activator of the ompF and ompC genes in Escherichia coli.

An in vivo approach was taken to assess whether the phosphorylated state of the transcription activator OmpR was affected by changes in the osmolarity of the growth medium or by mutations in envZ, the gene encoding the inner membrane histidine kinase that phosphorylates OmpR. We present results that support the view that increased phosphorylation of OmpR is correlated with enhanced expression of ompC. The in vivo phosphorylation approach was also used to show that OmpR can be phosphorylated in an envZ null strain. This result indicates that phosphorylation cross talk can occur in vivo between OmpR and a kinase(s) that is functionally homologous to envZ.

Bacterial Outer Membrane Proteins↗
PubMed1989

Cystic teratoma of the mesentery.

Cystic teratoma is an unusual cause of mesenteric/omental cysts. A case of cystic teratoma of the mesentery in a 5-year-old girl is reported.

Child, Preschool↗
PubMed1989

Phosphorylation of OmpR by the osmosensor EnvZ modulates expression of the ompF and ompC genes in Escherichia coli.

EnvZ and OmpR, the regulatory proteins for ompF and ompC expression in Escherichia coli, belong to a modulator-effector family of regulatory proteins which are essential for the response to environmental signals. We show that the soluble cytoplasmic domain of the transmembrane modulator protein EnvZ is phosphorylated in vitro by [gamma-32P]-ATP. We also demonstrate that the phosphate group can, in turn, be transferred to the transcription activator protein OmpR. The pH stability properties of the phosphate groups linked to EnvZ indicate that this molecule contains histidyl phosphate. The invariant His-243 of EnvZ corresponds to the phosphorylated His-48 of the chemotactic modulator protein CheA. Substitution of His-243 with valine produces an EnvZ that is refractory to phosphorylation and can no longer catalyze the transfer of phosphate to OmpR. Furthermore, in a delta envZ strain of E. coli, containing the envZ Val-243 plasmid, ompC expression is elevated 7-fold relative to that found in cells carrying the wild-type envZ plasmid. Based on these results we propose a model in which the phosphorylated state of OmpR modulates the expression of the ompF and ompC genes.

Bacterial Outer Membrane Proteins↗
PubMed1989

DNA-binding properties of the transcription activator (OmpR) for the upstream sequences of ompF in Escherichia coli are altered by envZ mutations and medium osmolarity.

Expression in Escherichia coli of the genes that encode the major outer membrane porin proteins (OmpF and OmpC) is regulated by the transcription activator protein OmpR and the receptorlike protein EnvZ, which is located in the inner membrane. Using synthesized oligonucleotide fragments containing the OmpR-binding site of ompF, we show that soluble extracts and partially purified OmpR derived from both the parent strain grown in nutrient broth plus 20% sucrose and the envZ11 strain grown in nutrient broth produced high-affinity DNA-binding activity, whereas soluble extracts from the parent strain grown in nutrient broth produced low-affinity binding. We also show that the soluble extracts from the envZ22(Am) strain grown in nutrient broth did not produce detectable bound forms of the ompF fragments, but low levels of DNA binding were detected with soluble extracts of the envZ22 strain grown in nutrient broth plus sucrose. In addition, the time course of the repression of OmpF synthesis produced by a shift to high-osmolarity growth medium was correlated with an increase in the DNA-binding affinity of soluble extracts to the ompF fragment. These results provide evidence that envZ function influences the DNA-binding activity of OmpR and suggest that high-affinity binding of OmpR to the upstream sequences of ompF is correlated with the repression of OmpF production.

Bacterial Outer Membrane Proteins↗
PubMed1989

[Conservative treatment of urethral stenosis in childhood].

During a 19 years period (1969-March 1988) 22 children presented for treatment of urethral strictures. The patients ranged in age from neonatal period to 13 years old, 20 of them were boys and 2 girls. We have found 3 congenital cases, 5 traumatic cases, 9 inflammatory (versus infections) cases and 5 iatrogenic cases. Diagnosis can be suspected from the history and physical examination (voided stream), and confirmed radiographically and endoscopically; we have employed urodynamics studies in 2 cases, and those have seem of much important the follow-up. Treatment is controversial between different authors. In ours series progressive dilatation procedure have been elective (retrograde swelling catheter versus anterograde swelling catheter with cytostomy), with 86.3% of good results.

Adolescent↗
PubMed1988

Purification, amino terminal analysis, and peptide mapping of proteins after in situ postelectrophoretic fluorescent labeling.

Proteins fractionated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis were stained in situ with either 5-(dimethylamino)-1-naphthalene sulfonyl chloride (dansyl chloride) or fluorescein isothiocyanate. This staining procedure can be carried out in less than 30 min without previous fixation of the proteins. It is not dependent on such factors as charge or molecular weight of the proteins and can detect 50 ng of protein in a 10-mm-wide gel slot. Fluorescent staining with dansyl chloride was used to localize proteins after electrophoresis for subsequent electroelution, amino terminal analysis, and peptide mapping. The electroelution can be carried out in less than 3 h with yields approaching 100%. The staining of only one strip of a preparative gel allowed the electroelution of proteins without covalent modification. For amino terminal analysis, identical results were obtained when the hydrolysis step was carried out after electroelution or directly in the gel pieces. The peptide mapping can be carried out with the proteins in solution (after electroelution) or directly in the gel pieces. The amino terminal and peptide mapping analysis of each protein in a mixture can be completed within 30 h from the beginning of the electrophoretic fractionation. The method appears to be applicable to a wide range of proteins showing very different biochemical properties.

Amino Acids↗
PubMed1988

Regulation of ompC and ompF expression in Escherichia coli in the absence of envZ.

The expression of the genes encoding the major outer membrane porin proteins OmpF and OmpC in Escherichia coli is regulated by ompR, which encodes the transcriptional activator protein OmpR, and envZ, which encodes a receptorlike protein located in the inner membrane. To examine the role of EnvZ in the expression of the osmoregulated porin genes, we analyzed the production of OmpF and OmpC in cells that lack envZ function. We show that EnvZ is required for the maximal production of OmpC in cells grown in minimal medium but is not essential for the efficient induction of OmpC that occurs during a shift to a high-osmolarity medium. In contrast, the production of OmpF in cells that lack envZ function was similar to that of the parent strain, whereas OmpF repression during a shift to a high-osmolarity medium was incomplete in the absence of EnvZ. These results are discussed in the context of the putative role of EnvZ in the expression of ompF and ompC.

Bacterial Outer Membrane Proteins↗
PubMed1987

[Transducing properties of lambda phagemids].

Transduction of genetic markers conferring drug-resistance with lambda phagemids has been studied in terms of the influence of various genetic conditions on the level of transduction. For this purpose, we have constructed and analysed a new series of phagemids - lambda gt::pBR322 and lambda gt::pUC19, in addition to earlier published phagemids. Experimental data indicate that the transduction frequency depends on the function of cI repressor and bacterial recA system, the plasmid orientation in the phagemid DNA, the selective marker used in an experiment and the concentration of the antibiotic in the medium.

Bacteriophage lambda↗
PubMed1986

[Psoriasis and chronic vitamin A poisoning].

A case of a five year old girl who during a treatment with high doses of A and D vitamins prescribed for a long time because "psoriasis" is showed. Clinical, radiologic and biochemical symptomatology of chronic poisoning by vitamin A is proved. Authors want to draw attention to dangers of long use of vitamin A.

Bone and Bones↗