Patrick Murray
Limerick Institute of Technology, Applied Sciuence, Faculty Member
Understanding how to address today’s global challenges is critical to improving corporate performance in terms of economic and environmental sustainability. In wastewater treatment systems, such an approach implies integrating efficient... more
Understanding how to address today’s global challenges is critical to improving corporate performance in terms of economic and environmental sustainability. In wastewater treatment systems, such an approach implies integrating efficient treatment technologies with aspects of the circular economy. In this business field, energy costs represent a large share of operating costs. This work discusses technological and management aspects leading to greater energy savings in Portuguese wastewater treatment companies. A mixed methodology, involving qualitative and quantitative aspects, for collecting and analysing data from wastewater treatment plants was used. The qualitative aspects consisted of a narrative analysis of the information available on reports and websites for 11 wastewater management companies in Portugal (e.g., technologies, treated wastewater volumes and operating costs) followed by a review of several international studies. The quantitative approach involved calculating th...
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Fungal biotechnology research has rapidly increased as a result of the growing awareness of sustainable development and the pressing need to explore eco-friendly options. In the nanotechnology field, silver nanoparticles (AgNPs) are... more
Fungal biotechnology research has rapidly increased as a result of the growing awareness of sustainable development and the pressing need to explore eco-friendly options. In the nanotechnology field, silver nanoparticles (AgNPs) are currently being studied for application in cancer therapy, tumour detection, drug delivery, and elsewhere. Therefore, synthesising nanoparticles (NPs) with low toxicity has become essential in the biomedical area. The fungus Chaetomium thermophilum (C. thermophilum) was here investigated—to the best of our knowledge, for the first time—for application in the production of AgNPs. Transmission electronic microscopy (TEM) images demonstrated a spherical AgNP shape, with an average size of 8.93 nm. Energy-dispersive X-ray spectrometry (EDX) confirmed the presence of elemental silver. A neutral red uptake (NRU) test evaluated the cytotoxicity of the AgNPs at different inhibitory concentrations (ICs). A half-maximal concentration (IC50 = 119.69 µg/mL) was used...
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β-glucans are a large class of complex polysaccharides with bioactive properties, including immune modulation. Natural sources of these compounds include yeast, oats, barley, mushrooms, and algae. Yeast is abundant in various processes,... more
β-glucans are a large class of complex polysaccharides with bioactive properties, including immune modulation. Natural sources of these compounds include yeast, oats, barley, mushrooms, and algae. Yeast is abundant in various processes, including fermentation, and they are often discarded as waste products. The production of biomolecules from waste resources is a growing trend worldwide with novel waste resources being constantly identified. Yeast-derived β-glucans may assist the host’s defence against infections by influencing neutrophil and macrophage inflammatory and antibacterial activities. β-glucans were long regarded as an essential anti-cancer therapy and were licensed in Japan as immune-adjuvant therapy for cancer in 1980 and new mechanisms of action of these molecules are constantly emerging. This paper outlines yeast β-glucans’ immune-modulatory and anti-cancer effects, production and extraction, and their availability in waste streams.
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Microalgae are at the start of the food chain, and many are known producers of a significant amount of lipids with essential fatty acids. However, the bioactivity of microalgal lipids for anti-inflammatory and antithrombotic activities... more
Microalgae are at the start of the food chain, and many are known producers of a significant amount of lipids with essential fatty acids. However, the bioactivity of microalgal lipids for anti-inflammatory and antithrombotic activities have rarely been investigated. Therefore, for a sustainable source of the above bioactive lipids, the present study was undertaken. The total lipids of microalga Chlorococcum sp., isolated from the Irish coast, were fractionated into neutral-, glyco-, and phospho-lipids, and were tested in vitro for their anti-inflammatory and antithrombotic activities. All tested lipid fractions showed strong anti-platelet-activating factor (PAF) and antithrombin activities in human platelets (half maximal inhibitory concentration (IC50) values ranging ~25–200 μg of lipid) with the highest activities in glyco- and phospho-lipid fractions. The structural analysis of the bioactive lipid fraction-2 revealed the presence of specific sulfoquinovosyl diacylglycerols (SQDG)...
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Fungi’s ability to convert organic materials into bioactive products offers environmentally friendly solutions for diverse industries. In the nanotechnology field, fungi metabolites have been explored for green nanoparticle synthesis.... more
Fungi’s ability to convert organic materials into bioactive products offers environmentally friendly solutions for diverse industries. In the nanotechnology field, fungi metabolites have been explored for green nanoparticle synthesis. Silver nanoparticle (AgNP) research has grown rapidly over recent years mainly due to the enhanced optical, antimicrobial and anticancer properties of AgNPs, which make them extremely useful in the biomedicine and biotechnology field. However, the biological synthesis mechanism is still not fully established. Therefore, this study aimed to evaluate the combined effect of time, temperature and pH variation in AgNP synthesis using three different fungi phyla (Ascomycota, Basidiomycota and Zygomycota) represented by six different fungi species: Cladophialophora bantiana (C. bantiana), Penicillium antarcticum (P. antarcticum), Trametes versicolor (T. versicolor), Trichoderma martiale (T. martiale), Umbelopsis isabellina (U. isabellina) and Bjerkandera adus...
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Lutein is particularly known to help maintain normal visual function by absorbing and attenuating the blue light that strikes the retina in our eyes. The effect of overexposure to blue light on our eyes due to the excessive use of... more
Lutein is particularly known to help maintain normal visual function by absorbing and attenuating the blue light that strikes the retina in our eyes. The effect of overexposure to blue light on our eyes due to the excessive use of electronic devices is becoming an issue of modern society due to insufficient dietary lutein consumption through our normal diet. There has, therefore, been an increasing demand for lutein-containing dietary supplements and also in the food industry for lutein supplementation in bakery products, infant formulas, dairy products, carbonated drinks, energy drinks, and juice concentrates. Although synthetic carotenoid dominates the market, there is a need for environmentally sustainable carotenoids including lutein production pathways to match increasing consumer demand for natural alternatives. Currently, marigold flowers are the predominant natural source of lutein. Microalgae can be a competitive sustainable alternative, which have higher growth rates and d...
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Microalgae have the potential to synthesize and accumulate lipids which contain high value fatty acids intended for nutrition and biodiesel applications. Nevertheless, lipid extraction methods for microalgae cells are not well established... more
Microalgae have the potential to synthesize and accumulate lipids which contain high value fatty acids intended for nutrition and biodiesel applications. Nevertheless, lipid extraction methods for microalgae cells are not well established and there is not a standard analytical methodology to extract fatty acids from lipid-producing microalgae. In this paper, current lipid extraction procedures employing organic solvents (chloroform/methanol, 2:1 and 1:2, v/v), sodium hypochlorite solution (NaClO), acid-catalysed hot-water extraction and the saponification process [2.5 M KOH/methanol (1:4, v/v)] have been evaluated with two species of microalgae with different types of cell walls. One is a marine diatom, Phaeodactylum tricornutum, and the other a freshwater green microalga, Haematococcus pluvialis. Lipids from all types of extracts were estimated gravimetrically and their fatty acids were quantified by a HPLC equipped with Q-TOF mass spectrometer. Results indicated significant differ...
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A β-N-acetylglucosaminidase produced by a novel fungal source, the moderately thermophilic aerobic ascomycete Talaromyces emersonii, was purified to apparent homogeneity. Submerged fermentation of T. emersonii, in liquid medium containing... more
A β-N-acetylglucosaminidase produced by a novel fungal source, the moderately thermophilic aerobic ascomycete Talaromyces emersonii, was purified to apparent homogeneity. Submerged fermentation of T. emersonii, in liquid medium containing algal fucoidan as the main carbon source, yielded significant amounts of extracellular N-acetylglucosaminidase activity. The N-acetylglucosaminidase present in the culture-supernatant was purified by hydrophobic interaction chromatography and preparative electrophoresis. The enzyme is a dimer with molecular weight and pI values of 140 and 3.85, respectively. Substrate specificity studies confirmed the glycan specificity of the enzyme for N-acetylglucosamine. Michaelis-Menten kinetics were observed during enzyme-catalyzed hydrolysis of the fluorescent substrate methylumbelliferyl-β-D-N-acetylglucosaminide at 50°C, pH 5.0 (Km value of 0.5 mM). The purified N-acetylglucosaminidase displayed activity over broad ranges of pH and temperature, yielding re...
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The thermostable enzyme systems produced by the thermophilic ascomycete fungus Talaromyces emersonii cultivated on various carbon sources were investigated for the production of high value products from sugar beet. A broad range of... more
The thermostable enzyme systems produced by the thermophilic ascomycete fungus Talaromyces emersonii cultivated on various carbon sources were investigated for the production of high value products from sugar beet. A broad range of enzymatic activities relevant to cellulose, ...
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The thermostable enzyme systems produced by the thermophilic ascomycete fungus Talaromyces emersonii cultivated on various carbon sources were investigated for the production of high value products from sugar beet. A broad range of... more
The thermostable enzyme systems produced by the thermophilic ascomycete fungus Talaromyces emersonii cultivated on various carbon sources were investigated for the production of high value products from sugar beet. A broad range of enzymatic activities relevant to cellulose, hemicellulose, and pectin hydrolysis were identified in T. emersonii culture filtrates. In hydrolysis experiments conducted at 71ºC, the enzyme cocktails generated sugar-rich syrups from untreated sugar beet plants. Maximal levels of sugar beet hydrolysis were obtained with T. emersonii enzyme cocktails induced with sorghum/ beet pulp (68%) and sugar beet plant (56%). The principle monosaccharides released were glucose, xylose, and arabinose with minor amounts of galactose and galacturonic acid. Northern analysis of RNA isolated from T. emersonii when sugar beet plants were used as the sole carbon inducing source showed that genes required for polysaccharide hydrolysis and five carbon monosaccharide metabolism w...
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Global concern over the depletion of fossil fuel reserves, and the detrimental impact that combustion of these materials has on the environment, is focusing attention on initiatives to create sustainable approaches for the production and... more
Global concern over the depletion of fossil fuel reserves, and the detrimental impact that combustion of these materials has on the environment, is focusing attention on initiatives to create sustainable approaches for the production and use of biofuels from various biomass substrates. The development of a low-cost, safe and eco-friendly process for the utilization of renewable resources to generate value-added products with biotechnological potential as well as robust microorganisms capable of efficient fermentation of all types of sugars are essential to underpin the economic production of biofuels from biomass feedstocks. Saccharomyces cerevisiae, the most established fermentation yeast used in large scale bioconversion strategies, does not however metabolise the pentose sugars, xylose and arabinose and bioengineering is required for introduction of efficient pentose metabolic pathways and pentose sugar transport proteins for bioconversion of these substrates. Our approach provided a basis for future experiments that may ultimately lead to the development of industrial S. cerevisiae strains engineered to express pentose metabolising proteins from thermophilic fungi living on decaying plant material and here we expand our original article and discuss the strategies implemented to improve pentose fermentation.
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A coupling of the plasmon on the surface of metal nanoparticles with an incident photon enhances a broad range of useful optical phenomena, such as resonant light scattering (RLS), surface plasmon resonance (SPR) or Raman scattering. Due... more
A coupling of the plasmon on the surface of metal nanoparticles with an incident photon enhances a broad range of useful optical phenomena, such as resonant light scattering (RLS), surface plasmon resonance (SPR) or Raman scattering. Due to these unique optical properties plasmonic nanostructures of different sizes and shapes have gained increasing popularity in areas such as cancer diagnosis, photothermal therapy as well as the imaging of living cells, detection of pathogens, biomolecules, metal ions, and the catalysis of various reactions in wet chemistry. This article reviews the current trends in the synthesis of plasmonic nanoparticles, particularly gold (AuNPs) and silver (AgNPs), using fungi as well as the proposed mechanisms for their mycofabrication. We provide an overview of the theoretical concepts of plasmonic nanoparticles which are sensitive electromagnetic responses that determine these nanoparticles applications.
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The first gene (α-gal1) encoding an extracellular α-D-galactosidase from the thermophilic fungus Talaromyces emersonii was cloned and characterized. The α gal1 gene consisted of an open reading frame of 1,792 base pairs interrupted by six... more
The first gene (α-gal1) encoding an extracellular α-D-galactosidase from the thermophilic fungus Talaromyces emersonii was cloned and characterized. The α gal1 gene consisted of an open reading frame of 1,792 base pairs interrupted by six introns that encoded a mature protein ...
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The first gene (alpha-gal1) encoding an extracellular alpha-Dgalactosidase from the thermophilic fungus Talaromyces emersonii was cloned and characterized. The alpha-gal1 gene consisted of an open reading frame of 1,792 base pairs... more
The first gene (alpha-gal1) encoding an extracellular alpha-Dgalactosidase from the thermophilic fungus Talaromyces emersonii was cloned and characterized. The alpha-gal1 gene consisted of an open reading frame of 1,792 base pairs interrupted by six introns that encoded a mature protein of 452 amino acids, including a 24 amino acid secretory signal sequence. The translated protein had highest identity with other fungal alpha-galactosidases belonging to glycosyl hydrolase family 27. The alpha-gal1 gene was overexpressed as a secretory protein with an N-terminal histidine tag in the methylotrophic yeast Pichia pastoris. Recombinant alpha-Gal1 was secreted into the culture medium as a monomeric glycoprotein with a maximal yield of 10.75 mg/l and purified to homogeneity using Hisbinding nickel-agarose affinity chromatography. The purified enzyme was maximally active at 70 degrees C, pH 4.5, and lost no activity over 10 days at 50 degrees C. alpha-Gal1 followed Michaelis-Menten kinetics ...
Research Interests: Technology, Kinetics, Gene expression, Biological Sciences, Sequence alignment, and 10 moreMicrobiology and Biotechnology, Affinity chromatography, Introns, Molecular cloning, Substrate Specificity, Amino Acid Sequence, Recombinant Proteins, Alpha-Galactosidase, Hydrogen-Ion Concentration, and *Hot Temperature
Mitochondrial malate dehydrogenase (m-MDH; EC 1.1.1.37), from mycelial extracts of the thermophilic, aerobic fungus Talaromyces emersonii, was purified to homogeneity by sequential hydrophobic interaction and biospecific affinity... more
Mitochondrial malate dehydrogenase (m-MDH; EC 1.1.1.37), from mycelial extracts of the thermophilic, aerobic fungus Talaromyces emersonii, was purified to homogeneity by sequential hydrophobic interaction and biospecific affinity chromatography steps. Native m-MDH was a dimer with an apparent monomer mass of 35 kDa and was most active at pH 7.5 and 52 degrees C in the oxaloacetate reductase direction. Substrate specificity and kinetic studies demonstrated the strict specificity of this enzyme, and its closer similarity to vertebrate m-MDHs than homologs from invertebrate or mesophilic fungal sources. The full-length m-MDH gene and its corresponding cDNA were cloned using degenerate primers derived from the N-terminal amino acid sequence of the native protein and multiple sequence alignments from conserved regions of other m-MDH genes. The m-MDH gene is the first oxidoreductase gene cloned from T. emersonii and is the first full-length m-MDH gene isolated from a filamentous fungal sp...
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Research Interests: Sequence alignment, Escherichia coli, Thermophilic, Glycosylation, European, and 9 moreMolecular cloning, Protein Secondary Structure Prediction, Molecular Characterization, Amino Acid Sequence, Base Sequence, Recombinant Proteins, Protein Binding, *Hot Temperature, and Biochemistry and cell biology
Research Interests: Microbiology, Multidisciplinary, Sequence Analysis, Phylogeny, Sequence alignment, and 9 moreMolecular cloning, Secondary Structure, Protein Secondary Structure Prediction, Substrate Specificity, Amino Acid Sequence, Hydrogen-Ion Concentration, *Hot Temperature, Gene expression profiling, and Intracellular
The thermophilic filamentous fungus Talaromyces emersonii secretes a variety of hydrolytic enzymes that are of interest for processing of biomass into fuel. Many carbohydrases have been isolated and characterized from this fungus, but no... more
The thermophilic filamentous fungus Talaromyces emersonii secretes a variety of hydrolytic enzymes that are of interest for processing of biomass into fuel. Many carbohydrases have been isolated and characterized from this fungus, but no studies had been performed on peptidases. In this study, two acid-acting endopeptidases were isolated and characterized from the culture filtrate of T. emersonii. One of these enzymes was identified as a member of the recently classified glutamic peptidase family and was subsequently named T. emersonii glutamic peptidase 1 (TGP1). The second enzyme was identified as an aspartyl peptidase (PEP1). TGP1 was cloned and sequenced and shown to exhibit 64 and 47% protein identity to peptidases from Aspergillus niger and Scytalidium lignocolum, respectively. Substrate profiling of 16 peptides determined that TGP1 has broad specificity with a preference for large residues in the P1 site, particularly Met, Gln, Phe, Lys, Glu, and small amino acids at P1' such as Ala, Gly, Ser, or Thr. This enzyme efficiently cleaves an internally quenched fluorescent substrate containing the zymogen activation sequence (k(cat)/K(m)=2 x 10(5) m(-1) s(-1)). Maximum hydrolysis occurs at pH 3.4 and 50 degrees C. The reaction is strongly inhibited by a transition state peptide analog, TA1 (K(i)=1.5 nM), as well as a portion of the propeptide sequence, PT1 (K(i)=32 nM). Ex vivo studies show that hyphal extension of T. emersonii in complex media is unaffected by the aspartyl peptidase inhibitor pepstatin but is inhibited by TA1 and PT1. This study provides insight into the functional role of the glutamic peptidase TGP1 for growth of T. emersonii.
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Recent sequencing of a number of fungal genomes has revealed the presence of multiple putative beta-glucosidases. Here, we report the cloning of two beta-glucosidase genes (bg1 and aven1), which have very different biological functions... more
Recent sequencing of a number of fungal genomes has revealed the presence of multiple putative beta-glucosidases. Here, we report the cloning of two beta-glucosidase genes (bg1 and aven1), which have very different biological functions and represent two of a number of beta-glucosidases from Talaromyces emersonii. The bg1 gene, encoding a putative intracellular beta-glucosidase, shows significant similarity to other fungal glucosidases from glycosyl hydrolase family 1, known to be involved in cellulose degradation. Solka floc, methyl-xylose, gentiobiose, beech wood xylan, and lactose induced expression of bg1, whereas glucose repressed expression. A second beta-glucosidase gene isolated from T. emersonii, aven1, encodes a putative avenacinase, an enzyme that deglucosylates the anti-fungal saponin, avenacin, rendering it less toxic to the fungus. This gene displays high homology with other fungal saponin-hydrolysing enzymes and beta-glucosidases within GH3. A putative secretory signal peptide of 21 amino acids was identified at the N-terminus of the predicted aven1 protein sequence suggesting that this enzyme is extracellular. Furthermore, T. emersonii cultivated on oat plant biomass was shown to deglucosylate avenacin. The presence of the avenacinase transcript was confirmed by RT-PCR on RNA extracted from mycelia grown in the presence of avenacin. The expression pattern of aven1 on various carbon sources was distinctly different from that of bg1. Only methyl-xylose and gentiobiose induced transcription of aven1. Gentiobiose induces synthesis of a number of cellulase genes by T. emersonii and it may be a possible candidate for the natural cellulase inducer observed in Penicillium purpurogenum. This work represents the first report of an avenacinase gene from a thermophilic, saprophytic fungal source, and suggests that this gene is not exclusive to plant pathogens.
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In this paper, we report new sequence data for secreted thermostable fungal enzymes from the un-sequenced xylanolytic filamentous fungus Talaromyces emersonii and reveal novel insights on the potential role of enzymes relevant as wheat... more
In this paper, we report new sequence data for secreted thermostable fungal enzymes from the un-sequenced xylanolytic filamentous fungus Talaromyces emersonii and reveal novel insights on the potential role of enzymes relevant as wheat dough improvers. The presence of known and de novo enzyme sequences were confirmed through NanoLC-ESI-MS/MS and resultant peptide sequences were identified using SWISS PROT databases. The de novo protein sequences were assigned identity based on homology to known fungal proteins. Other proteins were assigned function based on the limited T. emersonii genome coverage. This approach allowed the identification of enzymes with relevance as wheat dough improvers. Rheological examination of wheat dough and wheat flour components treated with the thermostable fungal enzyme cocktail revealed structural alterations that can be extrapolated to the baking process. Thermoactive amylolytic, xylanolytic, glucanolytic, proteolytic and lipolytic enzyme activities were observed. Previously characterized T. emersonii enzymes present included; β-glucosidase, xylan-1,4-β-xyloxidase, acetylxylan esterase, acid trehalase, avenacinase, cellobiohydrolase and endo-glucanase. De novo sequence analysis confirmed peptides as being; α-glucosidase, endo-1,4-β-xylanase, endo-arabinase, endo-glucanase, exo-β-1,3-glucanase, glucanase/cellulase, endopeptidase and lipase/acylhydrolase. Rheology tests using wheat dough and fractioned wheat flour components in conjunction with T. emersonii enzymes show the role of these novel biocatalysts in altering properties of wheat substrates. Enzyme treated wheat flour fractions showed the effects of particular enzymes on appropriate substrates. This proteomic approach combined with rheological characterization is the first such report to the authors' knowledge.
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Research Interests: Genetics, Microbiology, Physiology, Zoology, Immunology, and 35 morePlant Biology, Transcription Regulation, Salt Stress, Abiotic Stress, Cell Culture, Stress response, Experimental Botany, Protein structure, Fisheries Sciences, Gene, Aspergillus niger, Hydrogen Peroxide, Aspergillus Nidulans, Molecular cloning, Protein Sequence Analysis, Molecular Characterization, Amino Acid Profile, Molecular Mass, Plant species, Veterinary Sciences, SIGNAL PEPTIDE, Multiple Sequence Alignment, Amino Acid Sequence, Base Sequence, Fruit Ripening, Open Reading Frame, Expression analysis, Binding Site, Genome Mapping, *Hot Temperature, Full Length Movies, Nucleotides, Biochemistry and cell biology, Molecular Biology Reports, and Jasmonic Acid
A gene (cbh2) encoding cellobiohydrolase II was isolated from the fungus Talaromyces emersonii by rapid amplification of cDNA ends techniques and the equivalent genomic sequence was subsequently cloned. This represents the first report of... more
A gene (cbh2) encoding cellobiohydrolase II was isolated from the fungus Talaromyces emersonii by rapid amplification of cDNA ends techniques and the equivalent genomic sequence was subsequently cloned. This represents the first report of a key component of the cellulase regulon from this organism. DNA sequencing revealed that cbh2 has an open reading frame of 1377 bp, which encodes a putative polypeptide of 459 amino acids, and is interrupted by seven introns. The deduced amino acid sequence revealed that cbh2 has a modular structure with a predicted molecular mass of 47 kDa and consisting of a fungal type carbohydrate binding module separated from a catalytic domain by a proline/serine/threonine rich linker region. The deduced protein is homologous to fungal cellobiohydrolases in Family 6A of the glycosyl hydrolases. Profiles of cbh2 expression in T. emersonii investigated by Northern blot analysis revealed that expression is regulated at the transcriptional level. Expression of the T. emersonii cbh2 gene is induced by cellulose, xylan, xylose, and gentiobiose and clearly repressed by glucose. Putative regulatory element consensus sequences have been identified in the upstream regulatory sequence of the cbh2 gene including the catabolite repressor element and the activator of cellulase expression (Ace) binding sites. High sequence identity (67%) between the catalytic domain of Cel 6A from Trichoderma reesei and the T. emersonii cbh2 gene product allowed structure prediction for the 3D model of the T. emersonii catalytic domain to be a variant of the classical TIM alpha/beta fold.
Research Interests: Protein Structure Prediction, Cellulase, Sequence alignment, Promoter, Molecular cloning, and 11 moreAmino Acid Profile, Molecular Mass, Protein Conformation, Genome sequence, Amino Acid Sequence, Base Sequence, Open Reading Frame, Expression analysis, Binding Site, Biochemistry and cell biology, and DNA sequence
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Research Interests: Genetics, Organic Chemistry, Technology, Kinetics, Polysaccharides, and 52 moreBiological Chemistry, Gene expression, Aspergillus, Neurospora crassa, Biological Sciences, Infection and immunity, Phylogeny, Cerebrospinal Fluid, Humans, Sequence alignment, Microbiology and Biotechnology, Escherichia coli, Sphingomonas, Cytochrome C, Physical sciences, Temperature, Phylogenetic analysis, Renewable Resources, Aspergillus niger, Free Energy, Enzyme, Temperature Dependence, CHEMICAL SCIENCES, Molecular cloning, Trypanosoma brucei, Specific Activity, Substrate Specificity, Oxidative phosphorylation, Applied Biochemistry, Heat Shock Proteins, Biochemical Genetics, Sleeping sickness, S100 protein family, Erysipelothrix, Osmotic Stress, Amino Acid Sequence, Base Sequence, Chemistry Biology, Membrane Protein, Culture Media, Recombinant Protein, Heterologous Expression, Recombinant Proteins, Ascomycota, Structural Similarity Index, Alpha-Galactosidase, Hydrogen-Ion Concentration, Geologic Sediments, Carbon Source, Intracellular Space, Biochemistry and cell biology, and Adenosine Triphosphate
This work reports the first investigation of Remersonia thermophila hemicellulosic hydrolytic enzyme production, with subsequent purification of an extracellular endo-β-1,4-xylanase (RtXyl) and its application in bread making. The... more
This work reports the first investigation of Remersonia thermophila hemicellulosic hydrolytic enzyme production, with subsequent purification of an extracellular endo-β-1,4-xylanase (RtXyl) and its application in bread making. The research describes RtXyl purification from sorghum-induced submerged liquid cultures of this moderately thermophilic, aerobic, ascomycete fungus. The purified enzyme is a single subunit protein with a molecular mass of 42 kDa and exhibits glycosyl hydrolase family-10-like activity over a broad pH and temperature range. Optimal activity was measured at pH 6.0 and 65 °C respectively, which is suitable for bread making applications. Substrate specificity studies revealed that RtXyl is purely xylanolytic with no side-activities against other plant polysaccharides. The RtXyl catalytic efficiency (K cat/K m) was highest with oats spelt xylan (810.90 mg mL(-1) s(-1)), wheat arabinoxylan (809.52 mg mL(-1) s(-1)) and beechwood xylan (417.40 mg mL(-1) s(-1)) with less efficiency towards insoluble oats spelt xylan (236.40 mg mL(-1) s(-1)). Hydrolysis products analysed by thin layer chromatography yielded a range of xylosaccharides, predominantly xylotriose and xylobiose. RtXyl application in a basic wheat bread recipe at low dosages (0.297 XU/g) showed its suitability to increase loaf volume by 8.0 % compared with the control bread. RtXyl increased loaf softness by 19.6 % while reducing bread staling by 20.4 % up to 4 days of storage.