Biomedical subjects

Eliora Z Ron

Publications and source records attributed to Eliora Z Ron.

At least 19 recordsLinked to original sources

PubMed2004

Two-dimensional reference map of Agrobacterium tumefaciens proteins.

Proteomics based on two-dimensional (2-D) gel electrophoresis of proteins followed by spot identification with mass spectrometry is a commonly used method for physiological studies. Physiological proteomics requires 2-D reference maps, on which most of the main proteins are identified. We present a reference map for the bacterial plant pathogen Agrobacterium tumefaciens proteins, which contains more than 300 entries with an isoelectric point (pI) between 4 and 7. The quantitative study of the proteins in the analytical window of the master gel demonstrated unique features, in comparison with other bacteria. In addition, a theoretical analysis of several protein parameters was performed and compared with the experimental results. A comparison of the theoretical molecular weight (MW) of the proteins and their theoretical pI with their vertical and horizontal migration distances, respectively, pointed out the existence of several proteins that strongly diverted from the graph trend-line. These proteins were clearly subjected to post-translational modifications, which changed their pI and/or MW. Additional support for post-translational modifications comes from the identification of multiple spots of the same gene products. Post-translational modifications appear to be more common than expected, at least for soluble proteins, as more than 10% of the proteins were associated with multiple spots.

Agrobacterium tumefaciens↗

PubMed2003

Bacterial type III secretion systems are ancient and evolved by multiple horizontal-transfer events.

Type III secretion systems (TTSS) are unique bacterial mechanisms that mediate elaborate interactions with their hosts. The fact that several of the TTSS proteins are closely related to flagellar export proteins has led to the suggestion that TTSS had evolved from flagella. Here we reconstruct the evolutionary history of four conserved type III secretion proteins and their phylogenetic relationships with flagellar paralogs. Our analysis indicates that the TTSS and the flagellar export mechanism share a common ancestor, but have evolved independently from one another. The suggestion that TTSS genes have evolved from genes encoding flagellar proteins is effectively refuted. A comparison of the species tree, as deduced from 16S rDNA sequences, to the protein phylogenetic trees has led to the identification of several major lateral transfer events involving clusters of TTSS genes. It is hypothesized that horizontal gene transfer has occurred much earlier and more frequently than previously inferred for TTSS genes and is, consequently, a major force shaping the evolution of species that harbor type III secretion systems.

Bacteria↗

PubMed2003

Multilocus sequence typing (MLST) of Escherichia coli O78 strains.

Strains of Escherichia coli serotype O78 are associated with many diseases, including invasive infections, in humans and farm animals. The clonal relationship between strains from different hosts is therefore important for assessing the risk of zoonotic infections. Here we propose a multilocus sequence typing scheme for E. coli, based on six housekeeping genes. Preliminary, but significant, results indicate that clonal division in E. coli O78 strains is host independent, and closely related clones reside in different hosts. There was a positive correlation between virulence and clonal origin.

Acyltransferases↗

PubMed2003

Optical imaging fiber-based live bacterial cell array biosensor.

A live cell array biosensor was fabricated by immobilizing bacterial cells on the face of an optical imaging fiber containing a high-density array of microwells. Each microwell accommodates a single bacterium that was genetically engineered to respond to a specific analyte. A genetically modified Escherichia coli strain, containing the lacZ reporter gene fused to the heavy metal-responsive gene promoter zntA, was used to fabricate a mercury biosensor. A plasmid carrying the gene coding for the enhanced cyan fluorescent protein (ECFP) was also introduced into this sensing strain to identify the cell locations in the array. Single cell lacZ expression was measured when the array was exposed to mercury and a response to 100nM Hg(2+) could be detected after a 1-h incubation time. The optical imaging fiber-based single bacterial cell array is a flexible and sensitive biosensor platform that can be used to monitor the expression of different reporter genes and accommodate a variety of sensing strains.

Adenosine Triphosphatases↗

PubMed2003

On-line and in situ biosensors for monitoring environmental pollution.

Efficient tools for on-line and in situ monitoring of environmental pollutants are required to provide early warning systems. In addition, such tools can contribute important information on the progress of various remediation treatments. One of the recently developed monitoring technologies involves the use of whole-cell biosensors. Such biosensors could be constructed to detect general toxicity or specific toxicity caused by one or more pollutants. Currently, a large spectrum of microbial biosensors have been developed that enable the monitoring of pollutants by measuring light, fluorescence, color or electric current. Electrochemical monitoring is of special interest for in situ measurements as it can be performed using simple, compact and mobile equipment and is easily adaptable for on-line measurements. Here we survey the potential application of electrochemical biosensors in monitoring of general toxicity as well as hydrocarbons and heavy metals.

Bacteria↗

PubMed2003

Virulence and the heat shock response.

The major adaptive response to elevation in temperature is the heat shock response that involves the induction of many proteins--called heat shock proteins. These include chaperones, proteases, alternative sigma factors and other regulatory and structural proteins. The heat shock response is also turned on by other stress conditions, such as oxidative stress or pH changes. Bacterial entry into the host organism involves a significant environmental change, which is expected to induce the heat shock response. Indeed, some of the heat shock proteins are themselves virulence factors while others affect pathogenesis indirectly, by increasing bacterial resistance to host defenses or regulating virulence genes. The cross talk between heat shock and virulence genes is discussed.

Animals↗

PubMed2003

A novel ColV plasmid encoding type IV pili.

Many septicaemic Escherichia coli strains harbour ColV virulence plasmids. This paper describes pO78V, a conjugative ColV plasmid from an avian pathogenic E. coli strain that encodes type IV pili in addition to other virulence-related genes and tetracycline resistance. Plasmid location of type IV pili genes was demonstrated using Southern hybridization and expression of the pili was demonstrated using RT-PCR and phage sensitivity assays. This is a first report of a ColV plasmid encoding type IV pili. Plasmid pO78V is a mosaic plasmid containing replicons and other genes typical to both IncI1 and IncFII groups. As type IV pili of Gram-negative bacteria are involved in several stages of infection, their presence on a ColV virulence plasmid could expand the repertoire of pathogenesis-related genes.

Animals↗

PubMed2003

Petroleum pollution bioremediation using water-insoluble uric acid as the nitrogen source.

The biodegradation of hydrocarbon pollutants in open systems is limited by the availability of a utilizable nitrogen source. This limitation can be overcome by using uric acid. Enrichment cultures grown on crude oil-uric acid media yielded mixed and pure cultures that degraded petroleum. In a simulated open system, uric acid bound to crude oil and was available for bacterial growth and petroleum biodegradation.

Acinetobacter↗

PubMed2002

Role of fibronectin in curli-mediated internalization.

Curli fibers of Escherichia coli mediate internalization of bacteria by eukaryotic cells. As curli fibers bind fibronectin with high affinity, the role of fibronectin in the uptake process was studied. The experiments presented here support the involvement of fibronectin in internalization of bacteria. Furthermore, a peptide containing the RGD motif, responsible for interaction of fibronectin with cellular integrins, can strongly inhibit curli-mediated internalization. The ability of curli fibers to bind fibronectin can therefore be linked to virulence.

Bacterial Proteins↗

PubMed2002

Amperometric quantification of total coliforms and specific detection of Escherichia coli.

The quantitative determination of total and fecal coliforms, as indicators of fecal pollution, is essential for water quality control. We developed a sensitive, inexpensive amperometric enzyme biosensor based on the electrochemical detection of beta-galactosidase activity, using p-amino-phenyl-beta-D-galactopyranoside as substrate, for determining the density of coliforms, represented by Escherichia coli and Klebsiella pneumoniae. The specific detection of E. coli was achieved using an antibody-coated electrode that specifically binds the target bacteria. Amperometric detection enabled the determination of 1000 colony-forming units/mL within 60-75 min. Preincubation for 5-6 h further increased the sensitivity more than 100-fold. The present experimental setup allowed the simultaneous analysis of up to eight samples, using disposable screen-printed electrodes.

Antibodies↗

PubMed2002

Protein aggregation in Escherichia coli: role of proteases.

Protein aggregation is involved in several human diseases, and presumed to be an important process in protein quality control. In bacteria, aggregation of proteins occurs during stress conditions, such as heat shock. We studied the protein aggregates of Escherichia coli during heat shock. Our results demonstrate that the concentration and diversity of proteins in the aggregates depend on the availability of proteases. Aggregates obtained from mutants in the Lon (La) protease contain three times more protein than wild-type aggregates and show the broadest protein diversity. The results support the assumption that protein aggregates are formed from partially unfolded proteins that were not refolded by chaperones or degraded by proteases.

ATP-Dependent Proteases↗

PubMed2002

Molecular characterization of a salt-tolerant bacterial community in the rice rhizosphere.

The diversity of salt-tolerant bacteria present in the rhizosphere of Oryza sativa was investigated. Fourteen bacterial strains, isolated after enrichment in nitrogen-free, semi-solid medium and showing tolerance to 3% NaCl, were analyzed by restriction patterns produced by amplified DNA coding for 16S rDNA (ARDRA) with enzymes Sau3AI, AluI and RsaI which showed that they were represented by 4 ARDRA types. Biodiversity among the 14 strains was also analyzed by the random amplified polymorphic DNA (RAPD) technique with a 10-mer primer. Partial nucleotide sequence of 16S rDNA assigned these clusters to Serratia marcescens, Pseudomonas aeruginosa, Alcaligenes xylosoxidans and Ochrobactrum anthropi. Notably, all four bacterial species are potential human pathogens that infect immunocompromised patients.

Alcaligenes↗

PubMed2002

Biosurfactants and oil bioremediation.

Oil pollution is an environmental problem of increasing importance. Hydrocarbon-degrading microorganisms, adapted to grow and thrive in oil-containing environments, have an important role in the biological treatment of this pollution. One of the limiting factors in this process is the bioavailability of many fractions of the oil. The hydrocarbon-degrading microorganisms produce biosurfactants of diverse chemical nature and molecular size. These surface-active materials increase the surface area of hydrophobic water-insoluble substrates and increase their bioavailability, thereby enhancing the growth of bacteria and the rate of bioremediation.

Bacteria↗

PubMed2002

In vivo aggregation of a single enzyme limits growth of Escherichia coli at elevated temperatures.

The formation of protein aggregates is associated with unfolding and denaturation of proteins. Recent studies have indicated that, in Escherichia coli, cellular proteins tend to aggregate when the bacteria are exposed to thermal stress. Here, we show that the aggregation of one single E. coli cytoplasmic protein limits growth at elevated temperatures in minimal media. Homoserine trans-succinylase (HTS), the first enzyme in the methionine biosynthetic pathway, aggregates at temperatures higher than 44 degrees C in vitro. Above this temperature, we can also observe in vivo aggregation that results in the complete disappearance of the enzyme from the soluble fraction. Moreover, reducing the in vivo level of HTS aggregation enables growth at non-permissive temperatures. This is the first demonstration of the physiological role of aggregation of a specific protein in the growth of wild-type bacteria.

Acyltransferases↗

PubMed2002

Structure--function studies of the recombinant protein bioemulsifier AlnA.

Acinetobacter radioresistens KA53 produces an extracellular bioemulsifier, referred to as alasan. The surface active component of alasan is a 35.77 kDa protein AlnA. Although AlnA and the Escherichia coli outer membrane protein A (OmpA) have a high amino acid sequence homology, E. coli OmpA has no emulsifying activity. Comparison of the amino acid sequences of AlnA and E. coli OmpA revealed four hydrophobic regions in AlnA that were absent in E. coli OmpA. Deletions and substitutions (with the homologous OmpA sequence) were constructed in each of the four hydrophobic regions of AlnA by site-directed polymerase chain reaction (PCR) mutagenesis, using the overlap PCR method. Analysis of the emulsifying activities of the mutated AlnA molecules demonstrated that all four hydrophobic regions were necessary for emulsifying activity. However, most of the inactive mutated proteins still adhered avidly to hexadecane. These findings indicate that in addition to binding to hydrocarbons, the protein emulsifier must form a specific structure on the surface of the hydrocarbon that prevents coalescence of oil droplets. This is the first structure-function study of a protein bioemulsifer.

Acinetobacter↗

PubMed2002

The active component of the bioemulsifier alasan from Acinetobacter radioresistens KA53 is an OmpA-like protein.

The bioemulsifier of Acinetobacter radioresistens KA53, referred to as alasan, is a high-molecular-weight complex of polysaccharide and protein. Recently, one of the alasan proteins, with an apparent molecular mass of 45 kDa, was purified and shown to constitute most of the emulsifying activity. The N-terminal sequence of the 45-kDa protein showed high homology to an OmpA-like protein from Acinetobacter spp. In the research described here the gene coding for the 45-kDa protein was cloned, sequenced, and expressed in Escherichia coli. Recombinant protein AlnA (35.77 kDa without the leader sequence) had an amino acid sequence homologous to that of E. coli OmpA and contained 70% of the specific (hydrocarbon-in-water) emulsifying activity of the native 45-kDa protein and 2.4 times that of the alasan complex. In addition to their emulsifying activity, both the native 45-kDa protein and the recombinant AlnA were highly effective in solubilizing phenanthrene, ca. 80 microg per mg of protein, corresponding to 15 to 19 molecules of phenanthrene per molecule of protein. E. coli OmpA had no significant emulsifying or phenanthrene-solubilizing activity. The production of a recombinant surface-active protein (emulsification and solubilization of hydrocarbons in water) from a defined gene makes possible for the first time structure-function studies of a bioemulsan.

Acinetobacter↗

PubMed2002

Heat shock proteome of Agrobacterium tumefaciens: evidence for new control systems.

The regulation of Agrobacterium tumefaciens heat shock genes involves a transcriptional activator (RpoH) and repressor elements (HrcA-CIRCE). Using proteome analysis and mutants in these control elements, we show that the heat shock induction of 32 (out of 56) heat shock proteins is independent of RpoH and HrcA. These results indicate the existence of additional regulatory factors in the A. tumefaciens heat shock response.

Agrobacterium tumefaciens↗

PubMed1999

Genetic and functional analysis of genes required for the post-modification of the polyketide antibiotic TA of Myxococcus xanthus.

The antibiotic TA of Myxococcus xanthus is a complex macrocyclic polyketide, produced through successive condensations of acetate by a type I PKS (polyketide synthase) mechanism. The genes encoding TA biosynthesis are clustered on a 36 kb DNA fragment, which has been cloned and analysed. The chemical structure of TA and the mechanism by which it is synthesized indicate the need for several post-modification steps, which are introduced into the carbon chain of the polyketide to form the final bioactive molecule. These include the addition of several carbon atoms originating from acetate carbonyl, three C-methylations, O-methylation and a specific hydroxylation. This paper reports the analysis of five genes which are involved in the post-modification of TA. Their functional analysis, by specific gene disruption, suggests that they may be essential for the production of the active antibiotic. The characteristics and organization of the genes suggest that they may be involved in the addition of the carbon atoms which arise from acetate.

Amino Acid Sequence↗