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Wyniki wyszukiwania dla: L-methionine biosynthesis pathway
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Identification and cloning of C. albicans SC5314 genes encoding L-methionine biosynthetic pathway enzymes.
Dane BadawczeEnzymes of fungal L-methionine biosynthetic pathway: homoserine O-acetyltransferase (Met2p), O-acetylhomoserine sulfhydrylase (Met15p) and cystathionine-γ-synthase (Str2p) could be exploited as molecular targets for antifungal chemotherapy. The goal of the study was to identify and clone genes encoding mentioned above enzymes. MET2, MET15 and STR2 genes...
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Inserts amplification for knockout cassette construction
Dane BadawczeThe goal of the study was to obtain optimal conditions for amplification of upstream and downstream genes fragments and introduce a cleavage site for restriction enzymes into them thanks to which it will be possible to clone fragments into a plasmid containing elements of the knockout cassette. The constructed cassettes will enable the removal of the...
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Antifungal Effect of Penicillamine Due to the Selective Targeting of L-Homoserine O-Acetyltransferase
PublikacjaDue to the apparent similarity of fungal and mammalian metabolic pathways, the number of established antifungal targets is low, and the identification of novel ones is highly desirable. The results of our studies, presented in this work, indicate that the fungal biosynthetic pathway of L-methionine, an amino acid essential for humans, seems to be an attractive perspective. The MET2 gene from Candida albicans encoding L-homoserine...
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Overproduction of CaMet15p native and His-tag versions.
Dane BadawczeEnzyme of fungal L-methionine biosynthetic pathway: O-acetylhomoserine sulfhydrylase (Met15p) could be exploited as molecular target for antifungal chemotherapy. The goal of the study was to obtain conditions optimal for protein production with the use of prokaryotic cells. The constructed expression plasmids, designed in three different versions: encoding...
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Overproduction of homoserine O-acetyltransferase (CaMet2p) native and His-tag versions.
Dane BadawczeEnzyme of fungal L-methionine biosynthetic pathway: homoserine O-acetyltransferase (Met2p) could be exploited as molecular target for antifungal chemotherapy. The goal of the study was to obtain conditions optimal for protein production with the use of prokaryotic cells. The constructed expression plasmids, designed in three different versions: encoding...
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Characterization of two aminotransferases from Candida albicans
PublikacjaAminoadipate aminotransferase (AmAA) is an enzyme of α-aminoadipate pathway (AAP) for l-lysine biosynthesis. AmAA may also participated in biosynthesis or degradation of aromatic amino acids and in d-tryptophan based pigment production. The AAP is unique for fungal microorganisms. Enzymes involved in this pathway have specific structures and properties. These features can be used as potential molecular markers. Enzymes catalyzing...
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Antifungal Activity of Homoaconitate and Homoisocitrate Analogs
PublikacjaThirteen structural analogs of two initial intermediates of the L-alpha-aminoadipate pathway of L-lysine biosynthesis in fungi have been designed and synthesized, including fluoro- and epoxy-derivatives of homoaconitate and homoisocitrate. Some of the obtainedcompounds exhibited at milimolar range moderate enzyme inhibitory properties against homoaconitase and/or homoisocitrate dehydrogenase of Candida albicans. The structural...
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Construction of glucosamine-6-phosphate synthase with an internal hexahistydyl fragment
PublikacjaL-Glutamine: D-fructose-6-phosphate aminotransferase (EC 2.6.1.16), known also as glucosamine-6-phosphate (GlcN-6-P) syntase, catalyzes the first committed step in the amino sugar biosynthetic pathway in prokaryotic and eukaryotic organisms. The enzyme carries out a complex reaction involving ammonia transfer and sugar phosphate isomerization. GlcN-6-P syntase is an important point of metabolic control of amino sugar biosynthesis...
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Heptaene macrolides biosynthesis by Streptomyces species
PublikacjaPolyene macrolides are one of the groups of secondary metabolites, generated by microorganisms belonging to the Streptomyces genus. These compounds, containing 3 - 7 conjugated double bonds systems in their molecules exhibit high antifungal activity against a broad spectrum of fungal pathogens with heptaenes demonstrating the highest antifungal potential. At the large extent, efficiency of biosynthesis of these natural products...
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Phenotypic consequences of the LYS4 gene disruption in Candida albicans
PublikacjaThe main scientific purpose of our studies was to verify the hypothesis that homoaconitase (HA) from Candida albicans, an enzyme catalyzing a second step of the α-aminoadipate pathway (AAP) of L-Lys biosynthesis may become a new target for antifungal chemotherapy. Previous studies indicated that the A. fumigatus mutant lacking the functional lysF gene, encoding HA, exhibited attenuated virulence in a low-dose mouse infection model...
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Consequences of lysine auxotrophy for Candida albicans adherence and biofilm formation
PublikacjaA number of factors are known to be involved in Candida albicans virulence, although biofilm development on the surfaces of indwelling medical devices is considered to promote superficial or systemic disease. Based on previously reported up-regulation of saccharopine and acetyllysine in biofilm cells and activation of the lysine biosynthesis/degradation pathway, we investigated...
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Synthesis of phosphono and phosphate derivatives of hydroxyimino-D-alditols as new potential antifungal agents
PublikacjaIn search for new effective we antyfungals we focus on two enzymes involved in biosynthesis of the fungal cell wall. The first enzyme is glucosamine-6-phosphate (GlmS), which catalyzes transformation of D-fructose-6-phosphate (Fru-6P) to D-glucosamine-6-phodphate (GlcN-6P) in the chitin biosynthesis pathway. The second enzyme is phosphomannose isomerase (PMI) repored to play a crucial role in biosynthesis of many mannosylated structures,...
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Enzymes of the lysine biosynthetic pathway as targets for antifungals ?
PublikacjaSystemic infections caused by human pathogenic fungi in immunocompromized patients continue to be one of the important clinical problems. Limited availability of safe and efficacious antifungal chemotherapeutics and emerging resistance to existing drugs stimulates search for novel molecular targets for antifungals. The α-aminoadipate pathway (AAP) of L-lysine biosynthesis is unique in fungi and thus has been so far considered...
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Investigation of interaction between xanthene dye rhodamine B and metagenomic-derivated methylthioadenosine phosphorylase RSFP
PublikacjaIn the previous study we found that the expression of the metagenomic - derived methylthioadenosine phosphorylase gene (rsfp) in E. coli cells and the presence of rhodamine B in the growth medium were crucial for the pink fluorescence of one E. coli colony from Antarctic soil metagenomic library (Cieśliński et al., 2009). In this study, rsfp gene expression was induced by an arabinose promoter in an LMG194 E. coli strain and the...
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Novel application of terminal restriction fragments length polymorphism method (t-RFLP) for identification of six clinically important Candida sp.
PublikacjaTerminal restriction fragments length polymorphism method (t-RFLP) is a modified classical RFLP method. The main difference is based on carrying out PCR using a fluorescently labeled primer or primers. The result of such modification is that after digestion of amplicons only terminal fragments of PCR product (containing fluorescent dye) are visualized. T-RFLP method has found application in environmental microbiology and epidemiology....
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Regulation of LPS assembly via controlled proteolysis and sensing of LPS stress in Escherichia coli
PublikacjaLipopolysaccharide (LPS) is a complex glycolipid, essential for the bacterial viability and along with phospholipids, it constitutes the major amphiphilic component of outer membrane (OM) in most of the Gram-negative bacteria, including Escherichia coli. LPS molecules confer an effective permeability barrier to the OM and play a crucial role in bacteria-environment and -host interactions. The synthesis and accumulation of this...
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Hypericum alpestre extract exhibits in vitro and in vivo anticancer properties by regulating the cellular antioxidant system and metabolic pathway of L‐arginine
PublikacjaConventional treatment methods are not effective enough to fight the rapid increase in cancer cases. The interest is increasing in the investigation of herbal sources for the development of new anticancer therapeutics. This study aims to investigate the antitumor capacity of Hypericum alpestre (H. alpestre) extract in vitro and in vivo, either alone or in combination with the inhibitors of the L‐arginine/polyamine/nitric oxide...
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Carvacrol as a Stimulant of the Expression of Key Genes of the Ginsenoside Biosynthesis Pathway and Its Effect on the Production of Ginseng Saponins in Panax quinquefolium Hairy Root Cultures
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Characterization of recombinant homocitrate synthase from Candida albicans
PublikacjaLYS21 and LYS22 genes from Candida albicans encoding isoforms of homocitrate synthase (HCS), an enzyme catalyzing the first committed step in the L-lysine biosynthetic pathway, were cloned and expressed as NoligoHistagged fusion proteins in Escherichia coli. The purified gene products revealed HCS activity, i.e. catalyzed the condensation of α-ketoglutarate with acetyl-coenzyme A to yield homocitrate. The recombinant enzymes were purified...
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Characterization of d-xylose reductase, XyrB, from Aspergillus niger
PublikacjaD-xylose reductase is a member of the aldo-keto reductase family, and is involved in D-xylose and L-arabinose conversion through the Pentose Catabolic Pathway (PCP) in fungi. In this study, we biochemically characterized a newly identified second D-xylose reductase (XyrB) from Aspergillus niger. This NADPH-dependent reductase is able to efficiently convert D-xylose and L-arabinose, and it has the highest affinity for these sugars...