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At least 415 records · Page 23Linked to original sources
PubMed2002

Prospects for generating new antibiotics.

A plethora of human pathogens are now resistant to all clinically significant antibiotics causing a crisis, in the treatment and management of infectious diseases, but also presenting a clear danger to future public health. If drug resistance is going to be tackled successfully, new antibiotics must be continually developed to counteract the processes of evolution and natural selection in these populations of pathogens. Despite the introduction of powerful new technologies such as high throughput screening platforms and combinatorial chemistry, natural products still offer structural diversity worthy of screening for biological activity. Functional genomics can revolutionise rational drug design providing new targets for antimicrobial drug discovery. The clusters of genes, encoding enzymes that form bio-synthetic pathways leading to the synthesis of many natural products including polyketides and non-ribosomal peptides, are amenable to modern genetic engineering. Repositioning, deleting and replacing genes in these biosynthetic clusters has resulted in the synthesis of many 'un-natural' natural products. This review examines the engineering of proteins involved in chain initiation on polyketide synthases culminating in the production at high yield of a biologically active erythromycin derivative.

Drug Resistance, Microbial↗
PubMed2000

Protecting drinking water: water quality testing and PHAST in South Africa.

The paper presents an innovative field-based programme that uses a simple total coliform test and the approach of PHAST (Participatory Hygiene And Sanitation Transformation) to help communities exploring possible water quality problems and actions that can be taken to address them. The Mvula Trust, a South African water and environmental sanitation NGO, has developed the programme. It is currently being tested throughout South Africa. The paper provides two case studies on its implementation in the field, and suggests ways in which the initiative can be improved in the future.

Colony Count, Microbial↗
PubMed2003

Bioremediation of chromium contaminated environments.

Bioremediation is the most promising and cost effective technology widely used nowadays to clean up both soils and wastewaters containing organic or inorganic contaminants. Discharge of chromium containing wastes has led to destruction of many agricultural lands and water bodies. Utilisation of chromium(Cr) reducing microbes and their products has enhanced the efficiency of the process of detoxification of Cr(VI) to Cr(III). This review focuses mainly on the current technologies prevalent for remediation like natural attenuation, anaerobic packed bed bioreactors (using live cells, Cr(VI) reductases or their byproducts) and use of engineered microorganisms. Treatment of wastewaters by biosorption or using biofilms and immobilized microbial cells are also discussed.

Adsorption↗
PubMed1988

Plasmids in the environment.

Bacterial plasmids existed in bacteria before the antibiotic era but their presence was brought into prominence by the use of antibiotics which selected for antibiotic resistant strains. Subsequently, the range of genes carried on plasmids was shown to extend far beyond those coding for antibiotic resistance. Any consideration of plasmids in the environment, therefore, must include all plasmids whether or not they are genetically linked with antibiotic resistance. Antibiotic resistant bacteria may be found in the environment either by contamination with excreta from man and animals in which the strains were selected, or by their selection within the environment by antibiotics synthesized in situ or reaching the environment in an undegraded form in sewage from man and animals, or from industry. Other agents, also contaminating the environment, exert a selective pressure such as heavy metals in industrial effluents which select for metal resistance. This paper reviews the incidences and role of plasmids in various habitats including natural waters, soil, pastures, farm wastes, and human sewage from both hospitalised and other populations. Aspects of plasmid ecology, their biological role, and the transmissibility of genetic material between bacteria within the environment are considered. Two recent studies in Bristol, UK, are reported. The first was a genetic study on Escherichia coli isolates from calf slurry. Various DNA probes were used to determine the extent of gene exchange between the various serotypes within the natural environment. The second was a preliminary study to determine the stability of a recombinant plasmid, in a wild strain of Escherichia coli of pig origin, after its release into a semi-contained farm situation. It is now recognized that plasmids are widely distributed in bacterial populations in terrestrial and aquatic environments. Many have been detected by their carriage of genes coding for antibiotic or heavy metal resistance. Others, mainly cryptic in nature, have been demonstrated by plasmid profile studies on isolates from various habitats. Plasmids were shown to be present in a relatively few bacteria deposited in culture collections prior to the antibiotic era. Subsequently, the increased prevalence of R plasmids in bacteria in most ecosystems were due mainly to the selective pressure imposed by the use of antibiotics. This pressure may have been exerted either in the environment in which the strains were found or elsewhere, the environment subsequently being contaminated by antibiotic resistant bacteria.(ABSTRACT TRUNCATED AT 400 WORDS)

Animals↗
PubMed2006

Identification of genome-scale metabolic network models using experimentally measured flux profiles.

Genome-scale metabolic network models can be reconstructed for well-characterized organisms using genomic annotation and literature information. However, there are many instances in which model predictions of metabolic fluxes are not entirely consistent with experimental data, indicating that the reactions in the model do not match the active reactions in the in vivo system. We introduce a method for determining the active reactions in a genome-scale metabolic network based on a limited number of experimentally measured fluxes. This method, called optimal metabolic network identification (OMNI), allows efficient identification of the set of reactions that results in the best agreement between in silico predicted and experimentally measured flux distributions. We applied the method to intracellular flux data for evolved Escherichia coli mutant strains with lower than predicted growth rates in order to identify reactions that act as flux bottlenecks in these strains. The expression of the genes corresponding to these bottleneck reactions was often found to be downregulated in the evolved strains relative to the wild-type strain. We also demonstrate the ability of the OMNI method to diagnose problems in E. coli strains engineered for metabolite overproduction that have not reached their predicted production potential. The OMNI method applied to flux data for evolved strains can be used to provide insights into mechanisms that limit the ability of microbial strains to evolve towards their predicted optimal growth phenotypes. When applied to industrial production strains, the OMNI method can also be used to suggest metabolic engineering strategies to improve byproduct secretion. In addition to these applications, the method should prove to be useful in general for reconstructing metabolic networks of ill-characterized microbial organisms based on limited amounts of experimental data.

Computational Biology↗
PubMed2001

Engineering novel biocatalytic routes for production of semisynthetic opiate drugs.

The morphine alkaloids and their semisynthetic derivatives provide a diverse range of important pharmaceutical drugs. Current production of semisynthetic opiate drugs is by chemical means from naturally occurring morphine, codeine and thebaine. Although various microbial transformations of morphine alkaloids have been identified since the 1960s, more recently there has been considerable effort devoted to engineering biocatalytic routes for producing these important compounds. Such biocatalytic routes are attractive, as they would provide an alternative to the chemical production processes which suffer from limited supply of precursors, often low yields and toxic wastes. The biotransformation of morphine and codeine to the potent analgesic hydromorphone and the mild analgesic/antitussive hydrocodone, respectively, by recombinant Escherichia coli has been demonstrated and the problems encountered when engineering such a system will be discussed.

17-Hydroxysteroid Dehydrogenases↗
PubMed2005

Refolding and purification of non-fusion HPT protein expressed in Escherichia coli as inclusion bodies.

The gene encoding hygromycin B phosphotransferase (hpt) is a widely used selectable marker in the production of genetically engineered crops. To facilitate the safety assessment of this protein, the non-fusion hpt expression plasmid was constructed and introduced into Escherichia coli to produce enough quantity of the HPT protein. High level expressed HPT was achieved but most of the expressed protein aggregated as inclusion bodies. The inclusion bodies were washed, separated from the cells, and solubilized by 0.3% Sarkosyl. The protein was renatured by dilution and dialysis, and then purified by anion-exchange chromatography. The activity is 8 U/mg protein and the purity is about 95%. Further studies showed that the microbially produced HPT protein had comparable molecular weight, immuno-reactivities, N-terminal amino acid sequences, and biological activities with those of the HPT produced by transgenic rice harboring hpt gene. All these results demonstrated the validity of utilizing the microbially produced HPT to assess the safety of the HPT protein produced in genetically engineered rice.

Amino Acid Sequence↗
PubMed2003

Enhanced mercury biosorption by bacterial cells with surface-displayed MerR.

The metalloregulatory protein MerR, which exhibits high affinity and selectivity toward mercury, was exploited for the construction of microbial biosorbents specific for mercury removal. Whole-cell sorbents were constructed with MerR genetically engineered onto the surface of Escherichia coli cells by using an ice nucleation protein anchor. The presence of surface-exposed MerR on the engineered strains enabled sixfold-higher Hg(2+) biosorption than that found in the wild-type JM109 cells. Hg(2+) binding via MerR was very specific, with no observable decline even in the presence of 100-fold excess Cd(2+) and Zn(2+). The Hg(2+) binding property of the whole-cell sorbents was also insensitive to different ionic strengths, pHs, and the presence of metal chelators. Since metalloregulatory proteins are currently available for a wide variety of toxic heavy metals, our results suggest that microbial biosorbents overexpressing metalloregulatory proteins may be used similarly for the cleanup of other important heavy metals.

Adsorption↗
PubMed1998

Human dendritic cell (DC)-based anti-infective therapy: engineering DCs to secrete functional IFN-gamma and IL-12.

An imbalance in the Th1- and Th2-type cytokine responses may allow certain microbes to modify the host response to favor their own persistence. We now show that infection/pulsing of human CD34+ peripheral blood hemopoietic progenitor cell-derived dendritic cells (DCs) with Leishmania donovani promastigotes, Histoplasma capsulatum, and Mycobacterium kansasii impairs the constitutive production of IL-12 from these cells. Thus, strategies aimed at modulating a dysregulated Th1/Th2 response to infection would be of great interest. To both augment the host immune response and deliver potent immunomodulatory cytokines such as IL-12 and IFN-gamma, our goal is to develop a therapeutic strategy using genetically modified, microbial Ag-pulsed DCs. Toward developing such immunotherapies, we used retrovirus-mediated somatic gene transfer techniques to engineer human DCs to secrete biologically active IL-12 and IFN-gamma. DCs pulsed with microbial antigens (e.g., leishmania and histoplasma Ags) were capable of inducing proliferative responses in autologous CD4+ lymphocytes. CD4+ lymphocytes cocultured with IL-12-transduced autologous DCs had enhanced Ag-specific proliferative responses compared with CD4+ lymphocytes cocultured with nontransduced or IFN-gamma- transduced DCs. In this cell culture model system we demonstrate that IL-12 has a negative effect on IL-4 secretion that is independent of its ability to induce IFN-gamma secretion. Taken together, these results indicate that IL-12-transduced DCs may be specifically suited in inducing or down-modulating Ag-specific Th1 or Th2 responses, respectively, and thus may be useful as adjunctive therapy in those intracellular infections in which a dominant Th1 response is critical for the resolution of infection.

3T3 Cells↗
PubMed2025

Engineering of xylose metabolic pathways in Rhodotorula toruloides for sustainable biomanufacturing.

The oleaginous yeast Rhodotorula toruloides is a promising microbial cell factory for the sustainable production of biofuels and value-added chemicals from renewable carbon sources. Unlike the conventional yeast Saccharomyces cerevisiae, R. toruloides can naturally metabolize xylose, the second most abundant sugar in lignocellulosic hydrolysates. However, its native xylose metabolism is inefficient, characterized by slow xylose uptake and accumulation of D-arabitol. Moreover, despite its phenotype, research on the enzymes involved in xylose metabolism has yet to reach a consensus. Therefore, this review provides a comprehensive analysis of the non-canonical xylose metabolism in R. toruloides, focusing on the properties of key enzymes involved in xylose metabolism. Native xylose reductase and xylitol dehydrogenase exhibit broad substrate promiscuity compared to their counterparts in the xylose-fermenting Scheffersomyces stipitis. Additionally, the absence of xylulokinase expression under xylose-utilizing conditions redirects metabolism toward D-arabitol accumulation. Consequently, D-arabitol dehydrogenases and ribulokinase play essential roles in the xylose metabolism of R. toruloides. These findings highlight the fundamental differences between R. toruloides xylose metabolism and the oxidoreductase pathways observed in other xylose-fermenting yeast, providing insights for metabolic engineering strategies to improve xylose utilization and enhance bioconversion of cellulosic hydrolysates to different bioproducts by R. toruloides.

Xylose↗
PubMed1992

Opportunities for the cellular approach in biomedical engineering.

This review is a commentary on recent, altered perspectives about biomedical engineering and its role in medicine. It is argued that, rather than being a peripheral specialty, medical engineering and engineering principles in general have a direct application to biochemical medicine and cell biology. A brief description is given of the cell as a compartmentalised reactor system, and the ways in which it is possible to replace lost or aberrant cell function. Specific topics are then covered to illustrate the general thesis. These are: polymers for cell mimicry, cell-surface interactions, biomolecule transport, cell transport phenomena, cell signalling, harnessing of cells for therapy and microbial interactions. These disparate subject areas have a common thread of interest for the biomedical engineer, and are presented here in a way which highlights key points of relevance for engineering. Though necessarily brief, the various descriptions in this review provide a film indication that a rigorous approach to the assessment, modelling and use of cells along sound engineering lines is vital for the future. It is concluded that, without this approach, our understanding of cell biology will remain semiquantitative and semiempirical.

Biological Transport↗
PubMed1999

[Establishment of a microbial assay for screening anti-human immunodeficiency virus type-1 protease inhibitors].

OBJECTIVE: This study was to establish a microbial assay of human immunodeficiency virus type-1 protease (HIV-1 PR) activity for screening anti-HIV PR inhibitors. METHODS: A 24 bp synthetic oligonucleotide fragment that encodes the HIV-1 PR recognition sequence was inserted into the tetr gene of pBR322 (mtetr). Escherichia coli containing HIV-1 PR expression vector-pPOLO was transformed with pACYC184M containing modified mtetr gene. The transformant could express both HIV-1 PR and the modified Tet protein. RESULTS: The growth of engineered E. coli was prevented in the presence of tetracycline because the resistance Tet protein was degraded by HIV-1 PR. However inhibition of the HIV-1 PR restored tetracycline resistance. 31 chemical synthetic compounds were tested by the microbial assay. CONCLUSIONS: A microbial assay method of HIV-1 PR activity was established through a engineered E. coli. 5 mumol/L saqunavir-a special HIV-1 PR inhibitor showed inhibitory effect on the engineered E. coli. That means this model could be used as a initial screening model for anti-HIV PR agents.

Escherichia coli↗
PubMed2004

Bacterial diversity and function of aerobic granules engineered in a sequencing batch reactor for phenol degradation.

Aerobic granules are self-immobilized aggregates of microorganisms and represent a relatively new form of cell immobilization developed for biological wastewater treatment. In this study, both culture-based and culture-independent techniques were used to investigate the bacterial diversity and function in aerobic phenol- degrading granules cultivated in a sequencing batch reactor. Denaturing gradient gel electrophoresis (DGGE) analysis of PCR-amplified 16S rRNA genes demonstrated a major shift in the microbial community as the seed sludge developed into granules. Culture isolation and DGGE assays confirmed the dominance of beta-Proteobacteria and high-G+C gram-positive bacteria in the phenol-degrading aerobic granules. Of the 10 phenol-degrading bacterial strains isolated from the granules, strains PG-01, PG-02, and PG-08 possessed 16S rRNA gene sequences that matched the partial sequences of dominant bands in the DGGE fingerprint belonging to the aerobic granules. The numerical dominance of strain PG-01 was confirmed by isolation, DGGE, and in situ hybridization with a strain-specific probe, and key physiological traits possessed by PG-01 that allowed it to outcompete and dominate other microorganisms within the granules were then identified. This strain could be regarded as a functionally dominant strain and may have contributed significantly to phenol degradation in the granules. On the other hand, strain PG-08 had low specific growth rate and low phenol degradation ability but showed a high propensity to autoaggregate. By analyzing the roles played by these two isolates within the aerobic granules, a functional model of the microbial community within the aerobic granules was proposed. This model has important implications for rationalizing the engineering of ecological systems.

Aerobiosis↗
PubMed1997

Engineering factors affecting the granulation of sludge in the UASB reactor.

The influence of inoculum sludge involving different microbial populations, different modes of stream flow and different flow rates in a bioreactor on the granulation of sludge was studied. In the granulation process there were four periods: formation of microbial floccus, formation of subcore, growth of subcore, and maturity of granules. The production of microfloccus could be attributed to acid-forming bacteria. The hydrodynamic momentum transfer caused by moving phases and the hydraulic shear effect are the crucial factors affecting the formation of the subcore. For this a new concept-the lowest limited flow rate-is proposed, i.e., the lowest flow rate to form an expanded sludge bed. A sufficient feed rate (higher than the lowest flow rate), an appropriate sludge load, and an equal distribution of feed, as well as a suitable alkalinity in the medium are the essential factors for the scaling-up and process-control of the UASB reactor.

Anaerobiosis↗
PubMed2026

Environmental Release of Genetically Intervened Microorganisms: Towards a New Narrative.

The deliberate release of genetically engineered microorganisms for environmental applications has remained largely blocked since the early days of recombinant DNA technology, when limited ecological knowledge, lack of success stories and public apprehension shaped a culture of caution and restrictive regulation. Despite profound advances in microbial ecology, synthetic biology and genetic design, current frameworks still rely on outdated assumptions and legacy regulations that equate engineered microbes with inherent danger and demand unrealistic forms of absolute containment. This review examines how laboratory-trained microorganisms exist on a continuum with naturally evolved life, and that their risks are neither categorically different nor greater. Rather than pursuing unachievable containment, governance should shift towards traceability, stewardship and long-term monitoring through genomic barcodes, digital twins and transparent oversight. The vision moves from domination and control to care and partnership recognizing engineered microbes as live amendments capable of restoring degraded ecosystems. Achieving this transformation requires new terminology, phased field-trial frameworks, improved scaling methods, and the integration of epistemological perspectives that emphasize reciprocity and coexistence with nature. Reframing biotechnology in this way could finally unlock the capacity of engineered microorganisms to contribute responsibly and effectively to planetary repair in an era of escalating environmental crises.

Microorganisms, Genetically-Modified↗
PubMed2006

Genomics for environmental microbiology.

The utilization of natural microbial diversity in biotechnology is hindered by our inability to culture the vast majority of microorganisms and the observation that laboratory engineered bacteria rarely function in the wild. It is now clear that an understanding of the community structure, function and evolution of bacteria in their natural environments is required to meet the promise of microbial biotechnology. To meet these new challenges, microbiologists are applying the tools of genomics and related high-throughput technologies to both cultured microbes and environmental samples. This work will lead to new views on ecosystems and biological function together with the biotechnology enabled by this science.

Environmental Microbiology↗
PubMed2001

Quantification of intracellular metabolites in Escherichia coli K12 using liquid chromatographic-electrospray ionization tandem mass spectrometric techniques.

The quantitative comprehension of microbial metabolic networks is a prerequisite for an efficient rational strain improvement ("metabolic engineering"). It is therefore necessary to accurately determine the concentration of a large number of reactants (i.e., metabolites, nucleotides, cofactors) in order to understand "in vivo" reaction kinetics. Quantification of intracellular concentrations of glycolytic intermediates and nucleotides in Escherichia coli K12 using a perchloric acid extraction and an LC-ESI-MS method was achieved. Intracellular metabolites (e.g., glucose 6-phosphate, fructose 1,6-bisphosphate, 6-phospho gluconate, acetyl-CoA, adenine nucleotides) were quantified under defined (glucose-limited steady-state) growth conditions. The method was verified by comparing the intracellular metabolite concentrations measured via LC-ESI-MS with enzymatic determinations. It is thus possible to identify and quantify more than 15 intracellular metabolites in parallel with a minimal amount of sample volume.

Acetyl Coenzyme A↗
PubMed2007

A rare sugar xylitol. Part I: the biochemistry and biosynthesis of xylitol.

The rare sugar xylitol is a five-carbon polyol (pentitol) that has beneficial health effects. Xylitol has global markets and, therefore, it represents an alternative to current dominant sweeteners. The research on microbial reduction of D-xylose to xylitol has been focused on metabolically engineered Saccharomycess cerevisiae and Candida strains. The Candida strains have an advantage over the metabolically engineered S. cerevisiae in terms of D-xylose uptake and maintenance of the intracellular redox balance. Due to the current industrial scale production of xylitol, it has become an inexpensive starting material for the production of other rare sugar. The first part of this mini-review concentrates on the biochemistry of xylitol biosynthesis and the problems related to intracellular redox balance.

Biotechnology↗