Robert Mach
Tu Wien, Biochemical Technology, Faculty Member
ABSTRACT Fecal microbial pollution is a major problem throughout the Danube River Basin, posing a threat to various types of water use, including drinking water production from river bank filtrates, water supply for agricultural and... more
ABSTRACT Fecal microbial pollution is a major problem throughout the Danube River Basin, posing a threat to various types of water use, including drinking water production from river bank filtrates, water supply for agricultural and industrial use, and the role of the river as a recreational space. Fecal microbial pollution is introduced into the river by point sources, such as discharges of treated or untreated sewage from human sources or livestock, and by nonpoint sources, such as urban and agricultural runoff. In addition, fecal input from wildlife may be of importance in specific regions. Despite huge efforts to improve wastewater management in the past decade, in many sections, the river and its tributaries exhibit very high levels of fecal microbial pollution. To assess microbiological water quality, indicators of fecal pollution are used as surrogates for the potential presence of intestinal pathogens. However, the standard indicators cannot provide any reliable information regarding the origin of fecal pollution, nor can their concentration levels be directly related to human health risks for many types of exposure and situations. The aim of this book chapter is to summarize the historical developments in microbiological water quality research and to reflect the most recent publicly available data on the fecal microbial pollution status of the Danube River. Moreover, the first results on fecal microbial source tracking by molecular biology methods are presented along with their applicability in river water quality monitoring, including the monitoring of riparian wells and alluvial groundwater resources. Finally, a discussion of the general state of water quality and public health is presented concerning (i) the current situation and potential limitations of the Water Framework Directive regarding the microbiological quality elements, (ii) further improvements regarding sampling and monitoring strategies, and (iii) the recently introduced concept of “integrated framework of fecal pollution monitoring and management” and expected further methodological developments in the context of the Danube watershed. Rapid progress in research and development is currently being made in the area of fecal microbial source tracking, pathogen detection, and health risk assessment, and these innovations are also likely to complement basic fecal pollution monitoring programs for river systems such as the Danube in the near future.
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Der filamentöse Pilz Trichoderma reesei ist von Natur aus fähig pflanzliche Biomasse abzubauen. Er sondert eine Reihe von hydrolytischen Enzymen in seine Umgebung ab, die das pflanzliche Material, das zum größten Teil aus Zellulose und... more
Der filamentöse Pilz Trichoderma reesei ist von Natur aus fähig pflanzliche Biomasse abzubauen. Er sondert eine Reihe von hydrolytischen Enzymen in seine Umgebung ab, die das pflanzliche Material, das zum größten Teil aus Zellulose und Hemizellulose besteht, in niedermolekulare Kohlehydrate zerlegen. Die Abbauprodukte stehen nun dem Pilz zur Nahrungsaufnahme zur Verfügung. Je nachdem, welche Kohlehydrate in der Umgebung vorhanden sind, werden die regulatorischen Schaltkreise des Pilzes daran angepasst. Damit gibt es auch ein bevorzugtes Substrat für die Zellulasen- und Hemizellulasenproduktion. Beispielsweise übt der Einfachzucker D-Glukose, der leicht zu verstoffwechseln ist, eine Katabolitrepression auf die hydrolytischen Enzyme aus. Die Aufnahme solcher Einfachzucker wird priorisiert und die Energie wird vor allem in Biomassebildung und Selbsterhaltung gesteckt. Das stellt besonders für die industrielle Enzymproduktion eine große Herausforderung dar. Um dieses unerwünschte Phänom...
Sustainability and circularity are currently two relevant drivers in the development and optimisation of industrial processes. This study assessed the use of electrodialysis (ED) to purify synthetic erythritol culture broth and for the... more
Sustainability and circularity are currently two relevant drivers in the development and optimisation of industrial processes. This study assessed the use of electrodialysis (ED) to purify synthetic erythritol culture broth and for the recovery of the salts in solution, for minimising the generation of waste by representing an efficient alternative to remove ions, ensuring their recovery process contributing to reaching cleaner standards in erythritol production. Removal and recovery of ions was evaluated for synthetic erythritol culture broth at three different levels of complexity using a stepwise voltage in the experimental settings. ED was demonstrated to be a potential technology removing between 91.7–99.0% of ions from the synthetic culture broth, with 49–54% current efficiency. Besides this, further recovery of ions into the concentrated fraction was accomplished. The anions and cations were recovered in a second fraction reaching concentration factors between 1.5 to 2.5 time...
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can produce up to 100 g/liter of extracellular proteins. The major and industrially relevant products are cellobiohydrolase I (CBHI) and the hemicellulase XYNI. The genes encoding both enzymes are transcriptionally activated by the... more
can produce up to 100 g/liter of extracellular proteins. The major and industrially relevant products are cellobiohydrolase I (CBHI) and the hemicellulase XYNI. The genes encoding both enzymes are transcriptionally activated by the regulatory protein Xyr1. The first 850 nucleotides of the promoter contain 14 Xyr1-binding sites (XBS), and 8 XBS are present in the promoter. Some of these XBS are arranged in tandem and others as inverted repeats. One such element, an inverted repeat, plays a crucial role in the inducibility of the promoter. We investigated the impact of the properties of such elements by shuffling them by insertion, exchange, deletion, and rearrangement of elements in both the and promoter. A promoter-reporter assay using the gene allowed us to measure changes in the promoter strength and inducibility. Most strikingly, we found that an inverted repeat of XBS causes an important increase in promoter strength and allows induction by xylan or wheat straw. Furthermore, evi...
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Sorbicillinoids are a diverse group of yellow secondary metabolites that are produced by a range of not closely related ascomycetes, including , , and . They share a similarity to the name-giving compound sorbicillin, a hexaketide.... more
Sorbicillinoids are a diverse group of yellow secondary metabolites that are produced by a range of not closely related ascomycetes, including , , and . They share a similarity to the name-giving compound sorbicillin, a hexaketide. Previously, a conserved gene cluster containing two polyketide synthases has been identified as the source of sorbicillin, and a model for the biosynthesis of sorbicillin in has been proposed. In this study, we deleted the major genes of interest of the cluster in , namely , , and . Sor1 is the homolog of SorA, which is the first polyketide synthase of the proposed biosynthesis pathway. Sor3 is a flavin adenine dinucleotide (FAD)-dependent monooxygenase, and its homolog in , SorC, was shown to oxidize sorbicillin and 2',3'-dihydrosorbicillin to sorbicillinol and 2',3'-dihydrosorbicillinol, respectively, . Sor4 is an FAD/flavin mononucleotide-containing dehydrogenase with an unknown function. We measured the amounts of synthesized sorbicill...
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Due to its capability to secrete large quantities of plant biomass degrading enzymes (PBDE), is widely applied for industrial purposes. In nature, expression of PBDE is efficiently regulated in this fungus. Several factors involved in... more
Due to its capability to secrete large quantities of plant biomass degrading enzymes (PBDE), is widely applied for industrial purposes. In nature, expression of PBDE is efficiently regulated in this fungus. Several factors involved in this regulatory network have been identified. However, most of them are transcription factors. Long noncoding RNAs (lncRNAs) emerged as common players acting on epigenetic or transcriptional regulation in several eukaryotic organisms. To date, no lncRNA has been described in filamentous fungi. A lncRNA termed was identified in QM9414. In this study, it was characterized and evidence for its regulatory impact on cellulase expression was provided. Interestingly, different versions of were identified in different strains (namely, QM6a, QM9414, and Rut-C30), varying in terms of RNA length. Remarkably, considerable longer variants of this lncRNA are present in hypercellulolytic strains compared to the wild-type strain QM6a. Based on these results, a correla...
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Salivary gland hytrosaviruses (SGHVs, family ) are non-occluded dsDNA viruses that are pathogenic to some dipterans. SGHVs primarily replicate in salivary glands (SG), thereby inducing overt salivary gland hypertrophy (SGH) symptoms in... more
Salivary gland hytrosaviruses (SGHVs, family ) are non-occluded dsDNA viruses that are pathogenic to some dipterans. SGHVs primarily replicate in salivary glands (SG), thereby inducing overt salivary gland hypertrophy (SGH) symptoms in their adult hosts. SGHV infection of non-SG tissues results in distinct pathobiologies, including reproductive dysfunctions in tsetse fly, (Diptera: Glossinidae) and house fly. Infection with the virus (GpSGHV) resulted in the collapse of several laboratory colonies, which hindered the implementation of area wide integrated pest management (AW-IPM) programs that had a sterile insect technique (SIT) component. Although the impact of GpSGHV infection has been studied in some detail in , the impact of the virus infection on other tsetse species remains largely unknown. In the current study, we assessed the susceptibility of six species (, , , and ) to GpSGHV infections, and the impact of the viral infection on the fly pupation rate, adult emergence, and ...
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We report a novel molecular assay, based on helicase-dependent amplification (HDA), for the detection of enterococci as markers for fecal pollution in water. This isothermal assay targets the same Enterococcus 23S rRNA gene region as the... more
We report a novel molecular assay, based on helicase-dependent amplification (HDA), for the detection of enterococci as markers for fecal pollution in water. This isothermal assay targets the same Enterococcus 23S rRNA gene region as the existing quantitative polymerase chain reaction (qPCR) assays of the US Environmental Protection Agency Methods 1611 and 1609 but can be entirely performed on a simple heating block. The developed Enterococcus HDA assay successfully discriminated 15 enterococcal from 15 non-enterococcal reference strains and reliably detected 48 environmental isolates of enterococci. The limit of detection was 25 target copies per reaction, only three times higher than that of qPCR. The applicability of the assay was tested on 30 environmental water sample DNA extracts, simulating a gradient of fecal pollution. Despite the use of simple instruments, the HDA results were consistent with those of the qPCR reference. Given this performance, we conclude that the develop...
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Over the past 15 years, pioneering interdisciplinary research has been performed on the microbiology of hydrogeologically well-defined alpine karst springs located in the Northern Calcareous Alps (NCA) of Austria. This article gives an... more
Over the past 15 years, pioneering interdisciplinary research has been performed on the microbiology of hydrogeologically well-defined alpine karst springs located in the Northern Calcareous Alps (NCA) of Austria. This article gives an overview on these activities and links them to other relevant research. Results from the NCA springs and comparable sites revealed that spring water harbors abundant natural microbial communities even in aquifers with high water residence times and the absence of immediate surface influence. Apparently, hydrogeology has a strong impact on the concentration and size of the observed microbes, and total cell counts (TCC) were suggested as a useful means for spring type classification. Measurement of microbial activities at the NCA springs revealed extremely low microbial growth rates in the base flow component of the studied spring waters and indicated the importance of biofilm-associated microbial activities in sediments and on rock surfaces. Based on g...
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The increasing emergence of multi-resistant bacteria in healthcare settings, in the community and in the environment represents a major health threat worldwide. In 2016, we started a pilot project to investigate antimicrobial resistance... more
The increasing emergence of multi-resistant bacteria in healthcare settings, in the community and in the environment represents a major health threat worldwide. In 2016, we started a pilot project to investigate antimicrobial resistance in surface water. Bacteria were enriched, cultivated on selective chromogenic media and species identification was carried out by MALDI-TOF analysis. From a river in southern Austria a methicillin resistant Staphylococcus aureus (MRSA) was isolated. Whole genome sequence analysis identified the isolate as ST8, spa type t008, SCCmecIV, PVL and ACME positive, which are main features of CA-MRSA USA300. Whole genome based cgMLST of the water isolate and comparison to 18 clinical MRSA USA300 isolates from the Austrian national reference laboratory for coagulase positive staphylococci originating from 2004, 2005 and 2016 and sequences of 146 USA300 isolates arbitrarily retrieved from the Sequence Read Archive revealed a close relatedness to a clinical isol...
Long noncoding RNAs (lncRNAs) are crucial players in epigenetic regulation. They were initially discovered in human, yet they emerged as common factors involved in a number of central cellular processes in several eukaryotes. For example,... more
Long noncoding RNAs (lncRNAs) are crucial players in epigenetic regulation. They were initially discovered in human, yet they emerged as common factors involved in a number of central cellular processes in several eukaryotes. For example, in the past decade, research on lncRNAs in yeast has steadily increased. Several examples of lncRNAs were described in Saccharomyces cerevisiae and Schizosaccharomyces pombe. Also, screenings for lncRNAs in ascomycetes were performed and, just recently, the first full characterization of a lncRNA was performed in the filamentous fungus Trichoderma reesei. In this review, we provide a broad overview about currently known fugal lncRNAs. We make an attempt to categorize them according to their functional context, regulatory strategies or special properties. Moreover, the potential of lncRNAs as a biotechnological tool is discussed.
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Numerous bacterial genetic markers are available for the molecular detection of human sources of fecal pollution in environmental waters. However, widespread application is hindered by a lack of knowledge regarding geographical stability,... more
Numerous bacterial genetic markers are available for the molecular detection of human sources of fecal pollution in environmental waters. However, widespread application is hindered by a lack of knowledge regarding geographical stability, limiting implementation to a small number of well-characterized regions. This study investigates the geographic distribution of five human-associated genetic markers (HF183/BFDrev, HF183/BacR287, BacHum-UCD, BacH, and Lachno2) in municipal wastewaters (raw and treated) from 29 urban and rural wastewater treatment plants (750 - 4,400,000 population equivalents) from 13 countries spanning six continents. In addition, genetic markers were tested against 280 human and non-human fecal samples from domesticated, agricultural and wild animal sources. Findings revealed that all genetic markers are present in consistently high concentrations in raw (median log10 7.2 - 8.0 marker equivalents (ME) 100 ml-1) and biologically treated wastewater samples (median ...
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Fungi can produce a wide range of chemical compounds via secondary metabolism. These compounds are of major interest because of their (potential) application in medicine and biotechnology and as a potential source for new therapeutic... more
Fungi can produce a wide range of chemical compounds via secondary metabolism. These compounds are of major interest because of their (potential) application in medicine and biotechnology and as a potential source for new therapeutic agents and drug leads. However, under laboratory conditions, most secondary metabolism genes remain silent. This circumstance is an obstacle for the production of known metabolites and the discovery of new secondary metabolites. In this study, we describe the dual role of the transcription factor Xylanase promoter binding protein 1 (Xpp1) in the regulation of both primary and secondary metabolism of Trichoderma reesei Xpp1 was previously described as a repressor of xylanases. Here, we provide data from an RNA-sequencing analysis suggesting that Xpp1 is an activator of primary metabolism. This finding is supported by our results from a Biolog assay determining the carbon source assimilation behavior of an xpp1 deletion strain. Furthermore, the role of Xp...
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The industrially used ascomycete Trichoderma reesei secretes a typical yellow pigment during cultivation, while other Trichoderma species do not. A comparative genomic analysis suggested that a putative secondary metabolism cluster,... more
The industrially used ascomycete Trichoderma reesei secretes a typical yellow pigment during cultivation, while other Trichoderma species do not. A comparative genomic analysis suggested that a putative secondary metabolism cluster, containing two polyketide-synthase encoding genes, is responsible for the yellow pigment synthesis. This cluster is conserved in a set of rather distantly related fungi, including Acremonium chrysogenum and Penicillium chrysogenum As an attempt to silence the cluster in T. reesei, two genes of the cluster encoding transcription factors were individually deleted. For a complete genetic proof-of-function the genes were re-inserted into the genomes of the respective deletion strains. The deletion of the first transcription factor (termed Yellow Pigment Regulator 1 (Ypr1)) resulted in the full abolishment of the yellow pigment formation and the expression of most genes of this cluster. A comparative HPLC analysis of supernatants of the ypr1 deletion and its ...
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ABSTRACT The applicability of next generation DNA sequencing (NGS) methods for water quality assessment has so far not been broadly investigated. This study set out to evaluate the potential of an NGS-based approach in a complex catchment... more
ABSTRACT The applicability of next generation DNA sequencing (NGS) methods for water quality assessment has so far not been broadly investigated. This study set out to evaluate the potential of an NGS-based approach in a complex catchment with importance for drinking water abstraction. In this multi-compartment investigation, total bacterial communities in water, faeces, soil, and sediment samples were investigated by 454 pyrosequencing of bacterial 16S rRNA gene amplicons to assess the capabilities of this NGS method for (i) the development and evaluation of environmental molecular diagnostics, (ii) direct screening of the bulk bacterial communities, and (iii) the detection of faecal pollution in water. Results indicate that NGS methods can highlight potential target populations for diagnostics and will prove useful for the evaluation of existing and the development of novel DNA-based detection methods in the field of water microbiology. The used approach allowed unveiling of dominant bacterial populations but failed to detect populations with low abundances such as faecal indicators in surface waters. In combination with metadata, NGS data will also allow the identification of drivers of bacterial community composition during water treatment and distribution, highlighting the power of this approach for monitoring of bacterial regrowth and contamination in technical systems.
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The ascomycete Trichoderma reesei is one of the most well studied cellulolytic microorganisms. This fungus is widely used in the biotechnology industry, mainly in the production of biofuels. Due to its importance, its genome was sequenced... more
The ascomycete Trichoderma reesei is one of the most well studied cellulolytic microorganisms. This fungus is widely used in the biotechnology industry, mainly in the production of biofuels. Due to its importance, its genome was sequenced in 2008, opening new avenues to study this microorganism. In this 'post-genomic' era, a transcriptomic and proteomic era has emerged. Here, we present an overview of new findings in the gene expression regulation network of T. reesei. We also discuss new rational strategies to obtain mutants that produce hydrolytic enzymes with a higher yield, using metabolic engineering. Finally, we present how synthetic biology strategies can be used to create engineered promoters to efficiently synthesize enzymes for biomass degradation to produce bioethanol.
Research Interests: Gene regulation, Carbon, Regulation, Transcription Factors, Biological Control, and 17 moreGene expression, Starvation, Biological Sciences, Protein-DNA interaction, Biocontrol, Gene, Trichoderma, Aspergillus niger, Aspergillus Nidulans, Molecular cloning, Gen, DNA binding, Chitinase, DNA binding proteins, Binding Site, Response Elements, and Thallophyta
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Research Interests: Carbon, Gene expression, Multidisciplinary, Applied, Stress response, and 15 moreApplied Environmental Microbiology, Osmotic pressure, Trichoderma Harzianum, Trichoderma, Aspergillus niger, Western blot, Botrytis cinerea, Chitinase, Low Temperature, Base Sequence, Cell Wall, Enzyme Linked Immunosorbent Assay, Carbon Source, Oligosaccharides, and Glucose Oxidase
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ABSTRACT Regulation of formation of the extracellular xylanase system of Trichoderma reesei QM 9414 during growth on xylan, cellulose, and replacement onto a number of soluble inducers was investigated by Northern analysis of xyn1 and... more
ABSTRACT Regulation of formation of the extracellular xylanase system of Trichoderma reesei QM 9414 during growth on xylan, cellulose, and replacement onto a number of soluble inducers was investigated by Northern analysis of xyn1 and xyn2 transcripts and by the use of the Escherichia coli hph (hygromycin B-phosphotransferase-encoding) gene as a reporter. Whereas the xyn1 promoter is active in the presence of xylan and xylose, and virtually silenced in the presence of glucose, the xyn2 promoter enables basal transcription at a low level, but is enhanced in the presence of xylan and xylobiose and also of sophorose or cellobiose. The respective regulatory nucleotide regions were localized on a 221-base pair fragment and a 55-base pair fragment of the xyn1 and xyn2 5'-upstream noncoding sequences, respectively. Electrophoretic mobility shift assays, using cell-free extracts, identified induction-specific protein-DNA complexes: one complex of high mobility was observed under basal, noninduced conditions (glucose) with xyn2, which was in part replaced by a slow-migrating complex upon induction by xylan or sophorose. Both complexes bound to a CCAAT box. With xyn1, the induced complex also binds to a CCAAT box, but this binding is not observed in the presence of the carbon catabolite repressor Cre1, which binds to a nearby located consensus motif.
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A plate confrontation experiment is commonly used to study the mechanism by which Trichoderma spp. antagonize and parasitize other fungi. Previous work with chitinase gene expression (ech42) during the precontact period of this process in... more
A plate confrontation experiment is commonly used to study the mechanism by which Trichoderma spp. antagonize and parasitize other fungi. Previous work with chitinase gene expression (ech42) during the precontact period of this process in which cellophane and dialysis membranes separated Trichoderma harzianum and its host Rhizoctonia solani resulted in essentially opposite results. Here, we show that cellophane membranes are permeable to proteins up to at least 90 kDa in size but that dialysis membranes are not. ech42 was expressed during the precontact stage of the confrontation between Trichoderma atroviride and its host only if the cellophane was placed between the two fungi. These results are consistent with enzyme diffusion from T. atroviride to R. solani generating the trigger of ech42 gene expression.
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As the introduction and promotion of dehydrating toilets progresses, the safety of handling and reuse of their biosolids remains a question. A detailed study to understand the storage conditions and the fate of selected faecal indicators... more
As the introduction and promotion of dehydrating toilets progresses, the safety of handling and reuse of their biosolids remains a question. A detailed study to understand the storage conditions and the fate of selected faecal indicators was conducted on four urine diverting dehydrating toilet units, using ash as a major additive, in Kathmandu Valley, Nepal. Presumptive Escherichia coli, total coliforms, enterococci and different fractions of Clostridium perfringens were investigated under field storage conditions. In addition, chemo-physical and chemical (carbon, nitrogen, phosphorous content) parameters were investigated. Observed temperature was low in all the four toilets with a median of 24.0 degrees C, which was in the same range as the ambient temperature. pH was below the desired range of >9 and moisture level was very high (>60%). No single factor of the studied chemo-physical and chemical parameters could be found by statistical analysis to have accounted for the reduction of the indicators in any of the toilets. By time series analysis of the investigated strata in the faecal heaps (n=96), the determined reduction rate showed increasing persistence characteristics for E. coli, coliforms and enterococci with respective average log(10) reduction of -0.4, -0.3 and -0.2 per month (p<0.001). No significant reduction was observed for the different fractions of C. perfringens determined for the non-pasteurised and pasteurised fraction at 60 degrees C and 85 degrees C. 72% of randomly selected and analysed samples (n=36) were found to contain helminthes eggs. The used 6 months storage time did not prove sufficient to reach appropriate safety levels for handling and reuse of the biosolids.
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The cellulolytic potential of the wild-type strain of Trichoderma reesei was compared to other members of Trichoderma sect. Longibrachiatum and Hypocrea spp. that have anamorphs referable to that section. There was high diversity even... more
The cellulolytic potential of the wild-type strain of Trichoderma reesei was compared to other members of Trichoderma sect. Longibrachiatum and Hypocrea spp. that have anamorphs referable to that section. There was high diversity even within the same species (as defined by morphological and macromolecular characters). Differences, where notable, were more pronounced for carboxymethyl-cellulase activity than for filter paper activity. High cellulase activities were observed for several strains of T. longibrachiatum and T. citrinoviride, whereas T. parceramosum formed only low levels of activity. Among the corresponding teleomorphs, most strains of H. schweinitzii were comparatively poor producers, whereas the highest percentage of high producers was found among H. jecorina isolates, and many strains were even more active than the parent T. reesei QM 6a. Immunoblot analysis of corresponding culture filtrates of various H. jecorina strains showed that the three major cellulase proteins (cellobiohydrolase I, cellobiohydrolase II, and endoglucanase I) were present in culture filtrates and their M(r) was identical to that of the respective T. reesei proteins. ELISA analysis demonstrated that these enzymes were also present in the same relative proportions in culture filtrates from H. jecorina and T. reesei. With the aid of primers, corresponding to conserved sequences in the cellobiohydrolase I-encoding gene cbh1, a fragment of this gene was amplified from selected strains of H. jecorina, T. reesei, T. longibrachiatum, T. citrinoviride, and H. schweinitzii. The fragments had the same size in all fungi. Cleavage of this fragment with Hhal produced a RFLP pattern which was identical in H. jecorina and T. reesei, but different in the other species. In the latter, the RFLP pattern was also species specific. These results provide support for a close genetic similarity of T. reesei and H. jecorina cellulases. In the latter, an ascomycetous model system for cellulase biosynthesis is now available. The results further indicate that other anamorphs of Trichoderma section Longibrachiatum are promising sources of high cellulase production.
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The ascomycete Trichoderma reesei is used for the production of plant cell wall-degrading enzymes in industrial scale. The interplay of the transactivator Xyr1 and the repressor Cre1 mainly regulates the expression of these enzymes.... more
The ascomycete Trichoderma reesei is used for the production of plant cell wall-degrading enzymes in industrial scale. The interplay of the transactivator Xyr1 and the repressor Cre1 mainly regulates the expression of these enzymes. During induc-ing conditions, such as in the presence of sophorose, the transcription of the two major cellulase-encoding genes, cbh1 and cbh2, is activated as well as the expression of xyr1. In the presence of D-glucose carbon catabolite repression mediated by Cre1 takes place and the expression of Xyr1 and the plant cell wall-degrading enzymes is down-regulated. In this study we compare the chromatin status of xyr1, cbh1, and cbh2 promoters in the wild-type strain and the Cre1-deficient strain Rut-C30. Chromatin rearrangement occurs in the xyr1 promoter during induction on sophorose. Chromatin opening and protein-DNA interactions in the xyr1 promoter were detected especially in a region located 0.9 kb upstream the translation start co-don, which bears several putative Cre1-binding sites and a CCAAT-box. Moreover, the xyr1 promoter is overall more acces-sible in a cre1-truncated background, no matter which carbon source is present. This makes the xyr1 regulatory sequence a good target for promoter engineering aiming at the enhancement of cellulase production.
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The activity and regulation of alpha-aminoadipate reductase in three Penicillium chrysogenum strains (Q176, D6/1014/A, and P2), producing different amounts of penicillin, were studied. The enzyme exhibited decreasing affinity for... more
The activity and regulation of alpha-aminoadipate reductase in three Penicillium chrysogenum strains (Q176, D6/1014/A, and P2), producing different amounts of penicillin, were studied. The enzyme exhibited decreasing affinity for alpha-aminoadipate with increasing capacity of the respective strain to produce penicillin. The enzyme from all three strains was inhibited by L-lysine, and the enzyme from the lowest producer, Q176, was least sensitive. Between pH 7.5 and 6.5, inhibition of alpha-aminoadipate reductase by L-lysine was pH dependent, being more pronounced at lower pH. The highest producer strain, P2, displayed the lowest alpha-aminoadipate reductase activity at pH 7.0. In Q176, the addition of 0.5-1 mM of exogenous lysine stimulated penicillin formation, whereas the same concentration was ineffective or inhibitory with strains D6/1014/A and P2. The addition of higher (up to 5 mM) lysine concentrations inhibited penicillin production in all three strains. In mutants of P. chrysogenum D6/1014/A, selected for resistance to 20 mM alpha-aminoadipate, highest penicillin production was observed in those strains whose alpha-aminoadipate reductase was most strongly inhibited by L-lysine. The results support the conclusion that the in vivo activity of alpha-aminoadipate reductase from superior penicillin producer strains of P. chrysogenum is more strongly inhibited by lysine, and that this is related to their ability to accumulate increased amounts of alpha-aminoadipate, and hence penicillin.
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Page 1. FJ Schwarz, U. Meyer (Hrsg), Optimierung des Futterwertes von Mais und Maisprodukten 39 Mykotoxine in Lebens-und Futtermittel: Entwicklung verbesserter Testmethoden Mycotoxins in food and feed: Developments of improved analytical... more
Page 1. FJ Schwarz, U. Meyer (Hrsg), Optimierung des Futterwertes von Mais und Maisprodukten 39 Mykotoxine in Lebens-und Futtermittel: Entwicklung verbesserter Testmethoden Mycotoxins in food and feed: Developments of improved analytical methods ...