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O Käppeli

Publications and source records attributed to O Käppeli.

At least 19 recordsLinked to original sources
PubMed2001

Mycotoxin detoxication of animal feed by different adsorbents.

The contamination of animal feed with mycotoxins represents a worldwide problem for farmers. These toxins originate from molds whose growth on living and stored plants is almost unavoidable particularly under moist conditions. Mycotoxin-containing feed can cause serious diseases in farm animals resulting in suffering and even death and thus can cause substantial economic losses. The most applied method for protecting animals against mycotoxicosis is the utilization of adsorbents mixed with the feed which are supposed to bind the mycotoxins efficiently in the gastro-intestinal tract. Aluminosilicates are the preferred adsorbents, followed by activated charcoal and special polymers. The efficiency of mycotoxin binders, however, differs considerably depending mainly on the chemical structure of both the adsorbent and the toxin. This review describes the most important types of adsorbents and the respective mechanisms of adsorption. Data of the in vitro and in vivo efficacy of detoxication are given.

Adsorption↗
PubMed1998

Safety analysis of contained low-hazard biotechnology applications.

A technical safety analysis has been performed on a containment-level-2 pilot plant in order to assess an upgrading of the existing facility, which should comply with good manufacturing practices. The results were obtained by employing the hazard and operability (HAZOP) assessment method and are discussed in the light of the appropriateness of this procedural tool for low-hazard biotechnology applications. The potential release of micro-organisms accounts only for a minor part of the hazardous consequences. However, in certain cases the release of a large or moderate amount of micro-organisms would not be immediately identified. Most of the actions required to avoid these consequences fall into the realm of operational procedures. As a major part of potential failures result from human errors, standard operating procedures play a prominent role when establishing the concept of safety management. The HAZOP assessment method was found to be adequate for the type of process under investigation. The results also may be used for the generation of checklists which, in most cases, are sufficient for routine safety assurance.

Animals↗
PubMed1997

Safe biotechnology. 8. Transport of infectious and biological materials. Working Party "Safety in Biotechnology" of the European Federation of Biotechnology.

The transport of infectious and biological material is regulated by a number of international organizations. This mini-review has been compiled to increase awareness within the scientific community of problems caused by differences in terminology (such as infectious materials/substances, biological products, diagnostic specimens, genetically modified microorganisms) and certain technical aspects of the main international guidelines, and to assist policy makers in the creation of harmonized guidelines. A list of relevant Internet resources has been compiled.

Animals↗
PubMed1997

The science and intricacy of environmental safety evaluations.

Safety issues in biotechnology are subject to considerable controversy in public debate. Public concern is focused in particular on the degree of uncertainty associated with inadvertent and unforeseeable impacts of the deliberate or accidental release of genetically engineered organisms into the environment. The objective of this article is to demonstrate that there is a factual, scientific basis to environmental safety issues, by means of which the real, case-specific hazards requiring serious consideration by scientists, representatives of regulatory agencies and the public can be distinguished from conjectural hazards and naturally occurring background processes.

Biotechnology↗
PubMed1996

Safe biotechnology. 7. Classification of microorganisms on the basis of hazard. Working Party "Safety in Biotechnology" of the European Federation Biotechnology.

The current systems for classifying human pathogens on the basis of hazard are well developed and their basic criteria are in general agreement one with another. Of more importance, the safety practices based on these classifications have generally been successful. They have enabled extensive research activities, medical practice and industrial production to be conducted on an ever-increasing scale, involving dangerous microorganisms (e.g. in vaccine production and treatment of infected patients) with a very low incidence of adverse effects on the workers involved and the general public. Although the EU has adopted a harmonised list of agents in groups 1-4 there is as yet no complete agreement among member states and individual microbiologists. The purpose of this paper is to present a historical survey and to discuss the current processes for identifying and classifying the hazards posed by the use of microorganisms in research and technology. This is essential in the design of appropriate methods of counteracting potential risks.

Bacteria↗
PubMed1992

Factors affecting homologous overexpression of the Saccharomyces cerevisiae lanosterol 14 alpha-demethylase gene.

The Saccharomyces cerevisiae Lanosterol 14 alpha-demethylase (14DM) gene was overexpressed in S. cerevisiae using promoter sequences of the highly expressed S. cerevisiae glyceraldehyde-3-phosphate dehydrogenase (TDH3) gene. To investigate factors affecting 14DM overproduction, the levels of 14DM-specific RNAs, apoprotein, and heme protein, respectively, were determined and the 14DM-specific RNA levels compared with the RNA levels originating from the endogenous TDH gene(s). The quantitative measurements revealed that the 14DM steady-state RNA levels reached were some three- to five-fold below the theoretically expected values. With a view towards further improving expression of the 14DM gene, the spacing between the TDH3 promoter and the AUG was adjusted precisely and to rule out possible toxic effects exerted by the 14DM protein, the TDH3 promoter was placed under galactose regulation by introducing an UASG segment. Furthermore, the effects of the gene copy number on 14DM overproduction were investigated. From the analysis of the improved expression constructs five conclusions could be reached: (1) expression from the native 14DM gene is comparable to the expression driven by the TDH3 promoter-14DM fusion construct on single copy plasmid vectors; (2) expression from the TDH3 promoter-14DM construct on single-copy vectors is nearly as efficient as expression from the corresponding endogenous TDH3 gene; (3) the gene copy number has an effect on the relative expression levels of the TDH3 promoter-14DM constructs; (4) the steady-state amounts of protein produced are very nearly proportional to gene dosage; and (5) protein toxicity does not have a major impact on 14DM production. The maximum yield of 14DM was in the order of 7% of the total yeast protein and the maximum production of functional 14DM heme protein appears to be limited by the availability of heme.

Base Sequence↗
PubMed1991

Biosurfactants from Bacillus licheniformis: structural analysis and characterization.

The surface-active compounds of the strain Bacillus licheniformis were isolated and their structure was elucidated. The high surface-active capacity of the crude extract was basically due to traces of long-chain saturated fatty acids, especially of palmitic and stearic acids, to a mixture of small amounts of hydrocarbons with chain lengths of 20 and 22 carbons, and mainly to appreciable amounts of four slightly different lipopeptides. The lipopeptides were found to be a mixture of four closely related compounds. The lipophilic part consisting of i-, n-C14 or i-, ai-C15 beta-OH fatty acids was linked to the hydrophilic peptide moiety, which contained seven amino acids (Glu, Asp, Val, three Leu and Ile) by a lactone linkage. Fifteen milligrams per litre of the purified lipopeptide product decreased the surface tension of water from 72 mN m-1 to 27 mN m-1, characterizing the product as a powerful surface-active agent that compares favourably to other (bio)surfactants. Antibiotic activity was demonstrated against bacteria and yeasts.

Amino Acid Sequence↗
PubMed1991

Optimization of Candida tropicalis cytochrome P450alk gene expression in Saccharomyces cerevisiae with continuous cultures.

The cytochrome P450alk gene (P45alk) from Candida tropicalis ATCC 750 was expressed in Saccharomyces cerevisiae GRF18 under control of the alcohol dehydrogenase I (ADHI) promoter. To achieve stable expression over long time periods, a 2-microns derived replicative and an integrative expression system were tested in continuous culture. The 2-microns derived replicative system could not be maintained in cells over high generation numbers. In continuous culture, the instability was more pronounced at high dilution rates (D) and high histidine concentration, for which the yeast is auxotrophic. The nature of the instability was probably due to a gene conversion event between the plasmid and the yeast chromosome. In contrast, the integrative expression system was stably maintained in cells over prolonged cultivation times. Since this work focused on the production of large quantities of P450 by heterologous expression in yeast over prolonged time periods, the integrant was used to optimize P450alk expression by varying continuous culture parameters. The P450alk expression was shown to be dependent on the D applied to the culture. The highest P450alk expression levels were obtained at high D, when cell metabolism was shifted to partial glucose oxidation, yielding ethanol as a major metabolite in the culture supernatant. In contrast, when glucose was completely oxidized at low D, the ADHI-dependent P450alk expression was reduced and followed by a corresponding decrease in heterologous protein.

Alcohol Dehydrogenase↗
PubMed1991

Hydrocarbon assimilation and biosurfactant production in Pseudomonas aeruginosa mutants.

We isolated transposon Tn5-GM-induced mutants of Pseudomonas aeruginosa PG201 that were unable to grow in minimal media containing hexadecane as a carbon source. Some of these mutants lacked extracellular rhamnolipids, as shown by measuring the surface and interfacial tensions of the cell culture supernatants. Furthermore, the concentrated culture media of the mutant strains were tested for the presence of rhamnolipids by thin-layer chromatography and for rhamnolipid activities, including hemolysis and growth inhibition of Bacillus subtilis. Mutant 65E12 was unable to produce extracellular rhamnolipids under any of the conditions tested, lacked the capacity to take up 14C-labeled hexadecane, and did not grow in media containing individual alkanes with chain lengths ranging from C12 to C19. However, growth on these alkanes and uptake of [14C]hexadecane were restored when small amounts of purified rhamnolipids were added to the cultures. Mutant 59C7 was unable to grow in media containing hexadecane, nor was it able to take up [14C]hexadecane. The addition of small amounts of rhamnolipids restored growth on alkanes and [14C]hexadecane uptake. In glucose-containing media, however, mutant 59C7 produced rhamnolipids at levels about twice as high as those of the wild-type strain. These results show that rhamnolipids play a major role in hexadecane uptake and utilization by P. aeruginosa.

Alkanes↗
PubMed1990

Lanosterol 14 alpha-demethylase-encoding gene: systematic analysis of homologous overexpression in Saccharomyces cerevisiae using strong yeast promoters.

The Saccharomyces cerevisiae 14DM gene, encoding cytochrome P450 lanosterol 14 alpha-demethylase (14DM), was overexpressed in various S. cerevisiae strains under the control of three strong heterologous yeast transcription promoters (pADC1, pGPD, pPHO5) and under the control of its own promoter. Striking, strain-specific differences in 14DM transcription and in 14DM contents have been observed. The relative abundances of 14DM-specific mRNA and protein derived from a series of different expression plasmids were compared. It was found that the inducible PHO5 promoter in combination with the JL745 host led to the highest expression levels. 14DM-specific RNA reached up to 2% of the total cellular mRNA in this strain and approx. 3% of the total soluble yeast-cell protein was determined to be 14DM by quantitative Western blotting. By comparing the abundances of the different fusion transcripts with the transcript originating from the corresponding endogenous gene from which the promoter was derived, it could be concluded that the expression levels of the different 14DM fusion genes were far below the theoretically attainable values.

Base Sequence↗
PubMed1989

Genetic transformation of the filamentous yeast, Trichosporon cutaneum, using dominant selection markers.

An efficient transformation system for the filamentous yeast, Trichosporon cutaneum, has been developed. Transformation was obtained with plasmids carrying either the Escherichia coli hygromycin B phosphotransferase-encoding gene (hph) or the Streptoalloteichus hindustanus phleomycin-resistance gene (ble), as dominant selection markers. Expression of both resistance-conferring genes was controlled by the gpd promoter and the trpC terminator, from Aspergillus nidulans. The transformation frequency was up to 500 colonies/micrograms of transforming DNA, using the ble gene, and up to 100 colonies/micrograms of transforming DNA, using the hph gene. Co-transformation frequencies using unselected DNA varied between 50 and 65%. The transforming DNA was found to consist of multiple tandem plasmid copies of high Mr. This polymeric structure, in nonselective media, was mitotically unstable, possibly indicating that it existed in an episomal state.

Aspergillus nidulans↗
PubMed1987

Characterization of a soluble carnitine acetyltransferase from Candida tropicalis.

Carnitine acetyltransferase was purified from the cytoplasmic fraction of Candida tropicalis grown on alkanes in continuous culture. By ion-exchange chromatography the enzyme was resolved in two fractions with the same specific activity of 80 U/mg. The molecular mass of both enzyme forms, determined by non-denaturing gradient gel electrophoresis, was 540 kDa. After SDS electrophoresis only one band of 64 kDa was detected indicating that both enzymes are oligomers each containing eight subunits. Isoelectric focusing in agarose under non-denaturing conditions demonstrated the presence of at least four different charged species in the pH range between 5.6 and 6.7. After isoelectric focusing in 9 M urea/1% Nonidet P-40 gels, both enzyme forms were resolved into four bands. Peptide mapping, performed by cyanogen bromide cleavage of polypeptides separated by denaturing isoelectric focusing followed by second-dimension SDS electrophoresis, revealed a very high degree of homology between these polypeptides. The presence of the octameric form of carnitine acetyltransferase already in the starting material was demonstrated by non-denaturing gradient gel electrophoresis and immunoblotting. Antibodies against carnitine acetyltransferase from C. tropicalis ATCC 32113 formed precipitation lines with extracts from several Candida species but not with extracts of Candida utilis, Candida ethanothermophilum and an another strain of C. tropicalis.

Acetyltransferases↗
PubMed1987

Regulation of gluconeogenic enzymes during the cell cycle of Saccharomyces cerevisiae growing in a chemostat.

Oscillation of the activities of gluconeogenic enzymes (malate dehydrogenase, phosphoenolpyruvate carboxykinase and fructose-1,6-bisphosphatase) was observed during the cell cycle of chemostat cultures of Saccharomyces cerevisiae. Since ethanol is released by the cells at the beginning of the division cycle, its effect on enzyme expression was determined. Pulsing ethanol to a synchronously dividing yeast culture led to a prolongation of the metabolically active phase as indicated by the course of oxygen uptake and carbon dioxide production rates (concomitant ethanol and glucose assimilation). Enzyme activities also remained elevated as long as ethanol was available to the cells. After a substrate shift from glucose to ethanol during cell division, ethanol was used without a lag phase and enzyme induction increased from the level reached at the point of the substrate change. The data confirmed that the small amount of ethanol produced when the cells begin active reproduction acts as an inducer of gluconeogenic enzymes.

Cell Cycle↗
PubMed1986

The distinction of different types of cytochromes P-450 from the yeasts Candida tropicalis and Saccharomyces uvarum.

The distinction between two types of cytochromes P-450 originating from microsomes of Candida tropicalis grown on glucose and on alkane was achieved. Criteria of differentiation between these two cytochrome P-450 forms were based on the characteristics of reduced carbon monoxide difference spectra, on substrate specificity, and on binding and inhibition kinetics of the fungistatic compound propiconazole. One cytochrome P-450 form catalyzed the 14 alpha-demethylation of lanosterol and bound propiconazole with an equimolar ratio. This form was present in microsomes from glucose-grown cells and shared similar characteristics with the cytochrome P-450 originating from Saccharomyces uvarum grown on the same carbon source. The other cytochrome P-450 form catalyzed the terminal hydroxylation of aliphatic hydrocarbons and showed a less specific binding ratio with propiconazole (10(3) mol propiconazole for 1 mol cytochrome P-450). This type of cytochrome P-450 was only present in the microsomes of C. tropicalis grown on alkane.

Candida↗
PubMed1986

Pilot plant production of rhamnolipid biosurfactant by Pseudomonas aeruginosa.

Rhamnolipid biosurfactants were continuously produced with Pseudomonas aeruginosa on the pilot plant scale. Production and downstream processing elaborated on the laboratory scale were adapted to the larger scale. Differences in performance resulting from the scale-up are discussed. A biosurfactant concentration of approximately 2.25 g liter-1 was achieved. The biosurfactant yield on glucose was 77 mg g-1 h-1, and the productivity was 147 mg liter-1 h-1, corresponding to a daily production of 80 g of biosurfactant. The first enrichment step consisted of an adsorption chromatography which was followed by an anion-exchange chromatography. The resulting product was 90% pure, and the overall recovery of active material was above 60% with the downstream processing used.

Centrifugation↗