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Microbial production of L-tryptophan has been reviewed with 172 references. The review includes different tryptophan producing microorganisms, their optimal cultural conditions, yields, assay and process of recovery. It also includes a discussion on the pathway of tryptophan biosynthesis and its regulation. Achievements in this regard made through genetic engineering have also been included.
The biosynthesis of trehalose has been previously shown to serve as an important osmoprotectant and stress protectant in Escherichia coli. Our results indicate that overproduction of trehalose (integrated lacI-Ptac-otsBA) above the level produced by the native regulatory system can be used to increase the growth of E. coli in M9-2% glucose medium at 37 degrees C to 41 degrees C and to increase growth at 37 degrees C in the presence of a variety of osmotic-stress agents (hexose sugars, inorganic salts, and pyruvate). Smaller improvements were noted with xylose and some fermentation products (ethanol and pyruvate). Based on these results, overproduction of trehalose may be a useful trait to include in biocatalysts engineered for commodity chemicals.
Modern biotechnology encompasses many technologies. The one that has generated the most public interest is that of genetic engineering. The field has advanced dramatically since the early 1970's, when concern was so high that a moratorium was declared on research using recombinant DNA. Today there are numerous products such as human growth hormone, interferon and monoclonal antibody diagnostic kits on the market. Despite the obvious benefits of these products, there are still concerns over the environmental application of genetically engineered organisms. Many countries have assessed their existing legislation to ascertain its adequacy to regulate such intended releases. Numerous international organizations are becoming involved as well, and optimistically, regulations that are being developed in various countries may be harmonized. These activities have identified several areas where more information is required, including microbial taxonomy and classification, pathogenicity in target and non-target species, environmental behavior and risk assessment processes. Consensus at this time appears to be that our predictive capabilities are still developing and that new organisms should be evaluated on a case-by-case basis.
Mesocosm studies using sub-Antarctic soil artificially contaminated with diesel or crude oil were conducted in Kerguelen Archipelago (49 degrees 21' S, 70 degrees 13' E) in an attempt to evaluate the potential of a bioremediation approach in high latitude environments. All mesocosms were sampled on a regular basis over six months period. Soils responded positively to temperature increase from 4 degrees C to 20 degrees C, and to the addition of a commercial oleophilic fertilizer containing N and P. Both factors increased the hydrocarbon-degrading microbial abundance and total petroleum hydrocarbons (TPH) degradation. In general, alkanes were faster degraded than polyaromatic hydrocarbons (PAHs). After 180 days, total alkane losses of both oils reached 77-95% whereas total PAHs never exceeded 80% with optimal conditions at 10 degrees C and fertilizer added. Detailed analysis of naphthalenes, dibenzothiophenes, phenanthrenes, and pyrenes showed a clear decrease of their degradation rate as a function of the size of the PAH molecules. During the experiment there was only a slight decrease in the toxicity, whereas the concentration of TPH decreased significantly during the same time. The most significant reduction in toxicity occurred at 4 degrees C. Therefore, bioremediation of hydrocarbon-contaminated sub-Antarctic soil appears to be feasible, and various engineering strategies, such as heating or amending the soil can accelerate hydrocarbon degradation. However, the residual toxicity of contaminated soil remained drastically high before the desired cleanup is complete and it can represent a limiting factor in the bioremediation of sub-Antarctic soil.
Genes encoding plant antibiotic peptides show expression patterns that are consistent with a defence role. Transgenic over-expression of defence peptide genes is potentially useful to engineer resistance of plants to relevant pathogens. Pathogen mutants that are sensitive to plant peptides in vitro have been obtained and a decrease of their virulence in planta has been observed, which is consistent with their hypothetical defence role. A similar approach has been followed to elucidate the potential direct anti-microbial role of hydrogen peroxide. Additionally, a scavenger of peroxynitrite has been used to investigate its involvement in plant defence.
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Laboratory technologists (22%) developed infections with Shigella sonnei. The isolates had the same antibiogram and pulse-field gel electrophoresis pattern as an unknown isolate handled by a laboratory student. Covering faucet handles with paper towels during hand washing in the laboratory was protective. No further cases occurred after the laboratory was cleaned with a phenolic agent and a handle-free faucet was installed.
The beta-propeller fold appears as a very fascinating architecture based on four-stranded antiparallel and twisted beta-sheets, radially arranged around a central tunnel. Similar to the alpha/beta-barrel (TIM-barrel) fold, the beta-propeller has a wide range of different functions, and is gaining substantial attention. Some proteins containing beta-propeller domains have been implicated in the pathogenesis of a variety of diseases such as cancer, Alzheimer, Huntington, arthritis, familial hypercholesterolemia, retinitis pigmentosa, osteogenesis, hypertension, and microbial and viral infections. This article reviews some aspects of 3D structure, amino acids sequence regularities, and biological functions of the proteins containing beta-propeller domains. Major emphasis has been laid on beta-propellers whose functions are associated to human diseases. Recent research efforts reported in the fields of protein engineering, drug design, and protein structure-function relationship studies, concerning the beta-propeller architecture, have also been discussed.
The paper summarizes the findings of the physicochemical and potency of potassium fluoride peroxohydrate (PFP-1), a new Russian disinfecting agent having a universal spectrum of antimicrobial effects. It also summarizes the results of long-term practical use for the routine and final disinfection of rooms, sanitary engineering equipment, glassware, medical instruments against both sanitary indicative microorganisms, and actual pathogens of disease in infective and somatic hospitals, tuberculous dispensaries, sanatoria, schools, kindergartens, military hospitals, maternity houses of Russian different cities, towns, and localities. The paper presents comparative data on the mechanism of action of various modifications of PFP on microbial spores, on the stability of agents and their solutions during storage, on the damaging activity on the materials of the processed objects, on toxicological and ecological risks. The results of comparative studies of the bactericidal and sporicidal activities of the modifications PFP-1, PFP-2, PFP-3 confirmed that it is necessary to develop a procedure for preparing modifications containing high levels of hydrogen peroxide in the dry preparation and that they can be obtained at the pilot unit developed to prepare PFP-1 modifications.
Natural steroidal estrogen hormones, e.g., estrone (E1), 17beta-estradiol (E2), estriol (E3), and 17alpha-estradiol (17alpha), are released by humans and livestock in the environment and are the most potent endocrine disrupters even at nanogram per liter levels. Published studies broadly conclude that conventional wastewater treatment is efficient in the removal of 17beta-estradiol (85-99%), but estrone removal is relatively poor (25-80%). The removal occurs mainly through sorption by sludge and subsequent biodegradation. The long solids retention time in wastewater treatment systems enhances estrogen removal due to longer exposure and the presence of a diverse microbial community, particularly nitrifiers. In spite of the treatment, the effluent from conventional biological wastewater treatment systems still contains estrogenic compounds at a level that may cause disruption of endocrine systems in some species. Advanced wastewater treatment systems such as membrane processes remove the estrogen compounds mainly through physical straining of particle-bound estrogens. Another major source, which accounts for 90% of the estrogen load, is animal manure from concentrated animal-feeding operations (CAFOs). Manure is not required to be treated in the United States as long as it is not discharged directly into water bodies. Thus, there is an urgent need to study the fate of animal-borne estrogens from these facilities into the environment. A number of studies have reported the feminization of male aquatic species in water bodies receiving the effluents from wastewater treatment plants (WWTPs) or surface runoff from fields amended with livestock manure and municipal biosolids. Estrogenicity monitoring studies have been conducted in more than 30 countries, and abundant research articles are now available in refereed journals. This review paper focuses on estrogen contributions by wastewater and livestock manure, their removal rate and mechanisms in an engineered system, and their transport and ultimate fate in an engineered system and the environment. The review aims to advance our understanding of fate, transport, and biodegradation of estrogen compounds and outlines some directions for future research.
We studied the changes in water quality and formation of biofilms occurring in a pilot-scale water distribution system with two generally used pipe materials: copper and plastic (polyethylene, PE). The formation of biofilms with time was analysed as the number of total bacteria, heterotrophic plate counts and the concentration of ATP in biofilms. At the end of the experiment (after 308 days), microbial community structure, viable biomass and gram-negative bacterial biomass were analysed via lipid biomarkers (phospholipid fatty acids and lipopolysaccharide 3-hydroxy fatty acids), and the numbers of virus-like particles and total bacteria were enumerated by SYBR Green I staining. The formation of biofilm was slower in copper pipes than in the PE pipes, but after 200 days there was no difference in microbial numbers between the pipe materials. Copper ion led to lower microbial numbers in water during the first 200 days, but thereafter there were no differences between the two pipe materials. The number of virus-like particles was lower in biofilms and in outlet water from the copper pipes than PE pipes. Pipe material influenced also the microbial and gram-negative bacterial community structure in biofilms and water.
Ultrafiltration is the pressure-driven process by which hemodialysis removes excess fluid from renal failure patients. Despite substantial improvements in hemodialysis technology, three significant problems related to ultrafiltration remain: ultrafiltration volume control, ultrafiltration rate control, and backfiltration. Ultrafiltration volume control is complicated by the effects of plasma protein adsorption, hematocrit, and coagulation parameters on membrane performance. Furthermore, previously developed equations relating the ultrafiltration rate and the transmembrane pressure are not applicable to high-flux dialyzers, high blood flow rates, and erythropoietin therapy. Regulation of the ultrafiltration rate to avoid hypotension, cramps and other intradialytic complications is complicated by inaccurate estimates of dry weight and patient-to-patient differences in vascular refilling rates. Continuous monitoring of circulating blood volume during hemodialysis may enable a better understanding of the role of blood volume in triggering intradialytic symptoms and allow determination of optimal ultrafiltration rate profiles for hemodialysis. Backfiltration can occur as a direct result of ultrafiltration control and results in transport of bacterial products from dialysate to blood. By examining these problems from an engineering perspective, the authors hope to clarify what can and cannot be prevented by understanding and manipulating the fluid dynamics of ultrafiltration.
New plasmids pML2.1 and pML4 were constructed for cloning the transcription regulatory regions. In the pML2.1 the structural part of chloramphenicol acetyltransferase gene of the pBR325 is under control of the lacUV5-promotor. Because the unique BamH1 cleavage site is in the joint region, one may use it for cloning transcription termination regions and selecting recombinant clones with the AprCms phenotype. As for the pML4, the foreign fragment integration is carried directly before the structural part of cat-gene and it is expressed only if the promotor regions are present. The plasmids were sequenced and their restriction maps were established. Small molecular weight (about 2,0 MDa, AprCmr) or only Apr intact genes and convenient disposition of many unique cleavage sites by restriction endonucleases make these plasmids useful for different genetic engineering experiments.
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The coding potential of the genome of E. coli K-12 includes YgfO and YicE, two members of the evolutionarily conserved NAT/NCS2 transporter family that are highly homologous to each other (45% residue identity) and closely related to UapA of Aspergillus nidulans, a most extensively studied microbial member of this family. YgfO and yicE were cloned from the genome, over-expressed extrachromosomally and assayed for uptake of [(3)H]xanthine and other nucleobases, in E. coli K-12, under conditions of negligible activity of the corresponding endogenous systems. Alternative, essentially equivalent functional versions of YgfO and YicE were engineered by C-terminal tagging with an epitope from the E. coli lactose permease and a biotin-acceptor domain from Klebsiella pneumoniae. Both YgfO and YicE were shown to be present in the plasma membrane of E. coli and function as specific, high-affinity transporters for xanthine (K(m) 4.2-4.6 microM for YgfO, or 2.9-3.8 microM for YicE), in a proton motive force-dependent manner; they display no detectable transport of uracil, hypoxanthine, or uric acid at external concentrations of up to 0.1 mM. Both YgfO and YicE are inefficient in recognizing uric acid or xanthine analogues modified at position 8 of the purine ring (8-methylxanthine, 8-azaxanthine, oxypurinol, allopurinol), which distinguishes them from their fungal homologues UapA and Xut1.
BACKGROUND: In patients with inflammatory bowel diseases, T-cell activation driven by microflora has been implicated as a mechanism causing clonal expansion and infiltration of CD4+ T cells in colonic lamina propria (LP). We explored a regulatory mechanism preventing infiltration of CD4+ T cells specific to a microbe-associated antigen in the gut. METHODS: SCID mice were reconstituted with CD4+ T cells specific to ovalbumin (OVA) and were orally administered with Escherichia coli engineered to produce OVA. RESULTS: OVA-specific CD4+ T cells (KJ1-26+) were recruited to colonic LP in an Ag-dependent manner, which was inhibited by adoptive transfer of naturally occurring CD4+CD25+ T (Treg) cells. KJ1-26+ T cells and Treg cells are localized preferentially to the colonic follicles that contain dendritic cells. In mice given Treg cells, LP CD4+ T cells showed a decrease in proliferative and interferon gamma response and an increase in transforming growth factor beta1 response to OVA stimulation. Treg cells inhibited both antigenic activation of effector CD4+ T cells and class II/CD80/CD86 up-regulation of dendritic cells. CONCLUSION: : Treg cells suppress recruitment of CD4+ T cells specific to a microbe-associated antigen to LP, which was associated with colocalization of effector CD4+ T cells and Treg cells in colonic follicles.
Protease inhibitors (PIs) are an important class of drugs for the treatment of HIV infection. However, in the course of treatment, resistant viral variants with reduced sensitivity to PIs often emerge and become a major obstacle to successful control of viral load. On the basis of a compound equipotently inhibiting HIV-1 and 2 proteases (PR), we have designed a pseudopeptide inhibitor, QF34, that efficiently inhibits a wide variety of PR variants. In order to analyze the potency of the inhibitor, we constructed PR species harboring the typical (signature) mutations that confer resistance to commercially available PIs. Kinetic analyses showed that these mutated PRs were inhibited up to 1,000-fold less efficiently by the clinically approved PIs. In contrast, all PR species were effectively inhibited by QF34. In a clinical study, we have monitored 30 HIV-positive patients in the Czech Republic undergoing highly active antiretroviral therapy, and have identified highly PI resistant variants. Kinetic analyses revealed that QF34 retained its subnanomolar potency against multi-drug resistant PR variants. X-ray crystallographic analysis and molecular modeling experiments explained the wide specificity of QF34: this inhibitor binds to the PR in an unusual manner, thus avoiding contact sites that are mutated upon resistance development, and the unusual binding mode and consequently the binding energy is therefore preserved in the complex with a resistant variant. These results suggest a promising route for the design of second-generation PIs that are active against a variety of resistant PR variants.
The fermentation-derived 16-membered and 14-membered macrolides have been equally productive sources of semi-synthetic derivatives which have significantly extended the utility of the macrolide class as important antibiotics. New derivatives, prepared by both chemical and biochemical methods, have exhibited a variety of improved features, such as an expanded antimicrobial spectrum, increased potency, greater efficacy, better oral bioavailability, extended chemical and metabolic stability, higher and more prolonged concentrations in tissues and fluids, lower and less frequent dosing, and/or diminished side-effects [302]. However, even more improvements are both achievable and necessary if problems such as resistance to existing antibiotics continue to rise [303, 304]. Newer semi-synthetic macrolides which satisfy these important needs should be anticipated as the contributions from new fields such as genetic engineering of macrolide-producing organisms and more powerful computational chemistry are combined with the more traditional disciplines of chemical synthesis, bioconversions, and screening fermentation broths.