Ribosomes are ribonucleoprotein complexes highly conserved across all domains of life. The size differences of ribosomal RNAs (rRNAs) can be mainly attributed to variable regions termed expansion segments (ESs) protruding out from the ribosomal surface. The ESs were found to be involved in a range of processes including ribosome biogenesis and maturation, translation, and co-translational protein modification. Here, we analyze the rRNAs of the yeasts from the Magnusiomyces/Saprochaete clade belonging to the basal lineages of the subphylum Saccharomycotina. We find that these yeasts are missing more than 400 nt from the 25S rRNA and 150 nt from the 18S rRNAs when compared to their canonical counterparts in Saccharomyces cerevisiae. The missing regions mostly map to ESs, thus representing a shift toward a minimal rRNA structure. Despite the structural changes in rRNAs, we did not identify dramatic alterations in the ribosomal protein inventories. We also show that the size-reduced rRNAs are not limited to the species of the Magnusiomyces/Saprochaete clade, indicating that the shortening of ESs happened independently in several other lineages of the subphylum Saccharomycotina.

• Klíčová slova
• Magnusiomyces, expansion segments, ribosomal RNA, ribosome, yeast,
• MeSH
• fylogeneze MeSH
• molekulární evoluce MeSH
• ribozomální proteiny genetika   MeSH
• ribozomy * metabolismus   genetika   MeSH
• RNA ribozomální 18S genetika   MeSH
• RNA ribozomální * genetika   MeSH
• Saccharomyces cerevisiae genetika   MeSH
• Saccharomycetales genetika   klasifikace   metabolismus   MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• ribozomální proteiny MeSH
• RNA ribozomální 18S MeSH
• RNA ribozomální * MeSH
• RNA, ribosomal, 25S MeSH Prohlížeč

The purpose was to investigate a simultaneous biodegradation of the recalcitrant monoazo dye Reactive Orange 16 (RO16) in a mixed culture consisting of a biofilm of Pleurotus ostreatus-colonizing polyamide carrier and a suspension of the yeast Candida zeylanoides to see their biological interactions and possible synergistic action during degradation. Decolorization in the mixed culture was more effective than in the fungal monoculture, the respective decolorizations reaching 87.5% and 70% on day 11. The proliferation of yeast was reduced compared with the C. zeylanoides monoculture but enabled the yeast to participate in decolorization. The interaction of P. ostreatus with the yeast resulted in a gradual decrease of fungal manganese-dependent peroxidase (MnP) and laccase activities. Gas chromatography-mass spectrometry (GC-MS) analysis of the degradation products brought evidence that P. ostreatus split the dye molecule asymmetrically to provide 4-(ethenylsulfonyl) benzene whose concentration was much decreased in the mixed culture suggesting its increased metabolization in the presence of the yeast. In contrast, C. zeylanoides split the azo bond symmetrically producing the metabolites 4-(ethenylsulfonyl) aniline and α-hydroxybenzenepropanoic acid. Those metabolites were rapidly degraded in the mixed culture. A novel aspect is represented by the evidence of a mutual cooperative action of the fungal and yeast microorganisms in the mixed culture resulting in rapid decolorization and degradation of the dye.

• MeSH
• azosloučeniny metabolismus   MeSH
• biodegradace MeSH
• biofilmy MeSH
• chemické látky znečišťující vodu metabolismus   MeSH
• fungální proteiny metabolismus   MeSH
• lakasa metabolismus   MeSH
• metabolické sítě a dráhy MeSH
• mikrobiální interakce MeSH
• peroxidasy metabolismus   MeSH
• Pleurotus růst a vývoj   metabolismus   MeSH
• Saccharomycetales růst a vývoj   metabolismus   MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• azosloučeniny MeSH
• chemické látky znečišťující vodu MeSH
• fungální proteiny MeSH
• lakasa MeSH
• manganese peroxidase MeSH Prohlížeč
• peroxidasy MeSH
• reactive orange 16 MeSH Prohlížeč

To take full advantage of recombinant Pichia pastoris (Komagataella phaffii) as a production system for heterologous proteins, the complex protein secretory process should be understood and optimised by circumventing bottlenecks. Typically, little or no attention has been paid to the fate of newly synthesised protein inside the cell, or its passage through the secretory pathway, and only the secreted product is measured. However, the system's productivity (i.e. specific production rate qp), includes productivity of secreted (qp,extra) plus intracellularly accumulated (qp,intra) protein. In bioreactor cultivations with P. pastoris producing penicillin G acylase, we studied the dynamics of product formation, i.e. both the specific product secretion (qp,extra) and product retention (qp,intra) as functions of time, as well as the kinetics, i.e. productivity in relation to specific growth rate (μ). Within the time course, we distinguished (I) an initial phase with constant productivities, where the majority of product accumulated inside the cells, and qp,extra, which depended on μ in a bell-shaped manner; (II) a transition phase, in which intracellular product accumulation reached a maximum and productivities (intracellular, extracellular, overall) were changing; (III) a new phase with constant productivities, where secretion prevailed over intracellular accumulation, qp,extra was linearly related to μ and was up to three times higher than in initial phase (I), while qp,intra decreased 4-6-fold. We show that stress caused by heterologous protein production induces cellular imbalance leading to a secretory bottleneck that ultimately reaches equilibrium. This understanding may help to develop cultivation strategies for improving protein secretion from P. pastoris.Key Points• A novel concept for industrial bioprocess development.• A Relationship between biomass growth and product formation in P. pastoris.• A Three (3) phases of protein production/secretion controlled by the AOX1-promoter.• A Proof of concept in production of industrially relevant penicillin G acylase.

• Klíčová slova
• Fedbatch bioreactor cultivation, Penicillin G acylase, Pichia pastoris, Process optimisation, Secretion of a heterologous protein, Specific rate of product formation,
• MeSH
• bakteriální proteiny genetika   metabolismus   MeSH
• biomasa MeSH
• bioreaktory MeSH
• extracelulární prostor metabolismus   MeSH
• intracelulární prostor metabolismus   MeSH
• kinetika MeSH
• penicilinamidasa genetika   metabolismus   MeSH
• promotorové oblasti (genetika) MeSH
• rekombinantní proteiny genetika   metabolismus   MeSH
• Saccharomycetales genetika   růst a vývoj   metabolismus   MeSH
• techniky vsádkové kultivace MeSH
• teoretické modely MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• bakteriální proteiny MeSH
• penicilinamidasa MeSH
• rekombinantní proteiny MeSH

There are only a few antifungal drugs used systemically in treatment, and invasive fungal infections that are resistant to these drugs are an emerging problem in health care. In this study, we performed a high-copy-number genomic DNA (gDNA) library screening to find and characterize genes that reduce susceptibility to amphotericin B, caspofungin, and voriconazole in Saccharomyces cerevisiae We identified the PDR16 and PMP3 genes for amphotericin B, the RMD9 and SWH1 genes for caspofungin, and the MRS3 and TRI1 genes for voriconazole. The deletion mutants for PDR16 and PMP3 were drug susceptible, but the other mutants had no apparent susceptibility. Quantitative-PCR analyses suggested that the corresponding drugs upregulated expression of the PDR16, PMP3, SWH1, and MRS3 genes. To further characterize these genes, we also profiled the global expression patterns of the cells after treatment with the antifungals and determined the genes and paths that were up- or downregulated. We also cloned Candida albicans homologs of the PDR16, PMP3, MRS3, and TRI1 genes and expressed them in S. cerevisiae Heterologous expression of Candida homologs also provided reduced drug susceptibility to the budding yeast cells. Our analyses suggest the involvement of new genes in antifungal drug resistance.

• Klíčová slova
• amphotericin B, antifungal agents, caspofungin, drug resistance, genomics, multidrug resistance, voriconazole,
• MeSH
• amfotericin B farmakologie   MeSH
• antifungální látky farmakologie   MeSH
• Candida albicans účinky léků   genetika   metabolismus   MeSH
• fungální léková rezistence genetika   MeSH
• kaspofungin farmakologie   MeSH
• mikrobiální testy citlivosti MeSH
• Saccharomyces cerevisiae - proteiny genetika   metabolismus   MeSH
• Saccharomyces cerevisiae účinky léků   genetika   metabolismus   MeSH
• Saccharomycetales účinky léků   genetika   metabolismus   MeSH
• vorikonazol farmakologie   MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• amfotericin B MeSH
• antifungální látky MeSH
• kaspofungin MeSH
• Saccharomyces cerevisiae - proteiny MeSH
• vorikonazol MeSH

Antibiotic-resistant bacteria have been observed with increasing frequency over the past decades, driving the search for new drugs and stimulating the interest in natural products sources. Endophytic fungi from medicinal plants represent a great source of novel bioactive compounds useful to pharmaceutical and agronomical purposes. Diaporthe terebinthifolii is an endophytic species isolated from Schinus terebinthifolius, a plant used in popular medicine for several health problems. The strain D. terebinthifolii LGMF907 was previously reported by our group to produce secondary metabolites with biological activity against phytopathogens. Based on these data, strain LGMF907 was chosen for bioprospecting against microorganisms of clinical importance and for characterization of major secondary metabolites. In this study, different culture conditions were evaluated and the biological activity of this strain was expanded. The crude extracts demonstrated high antibacterial activity against Escherichia coli, Micrococcus luteus, Saccharomyces cerevisiae, methicillin-sensitive Staphylococcus aureus, and methicillin-resistant S. aureus. The compounds diaporthin and orthosporin were characterized and also showed activity against the clinical microorganisms evaluated. This study discloses the first isolation of diaporthin and orthosporin from D. terebinthifolii, and revealed the potential of this endophytic fungus to produce secondary metabolites with antimicrobial activity.

• Klíčová slova
• Diaporthe terebinthifolii, Diaporthin, Endophyte, Orthosporin, Schinus terebinthifolius Raddi, Secondary metabolites,
• MeSH
• antiinfekční látky chemie   izolace a purifikace   farmakologie   MeSH
• Bacteria účinky léků   MeSH
• bioprospekting * MeSH
• endofyty chemie   metabolismus   MeSH
• Escherichia coli účinky léků   MeSH
• fermentace MeSH
• kultivační média MeSH
• methicilin rezistentní Staphylococcus aureus MeSH
• mikrobiální testy citlivosti MeSH
• molekulární struktura MeSH
• Mycobacterium účinky léků   MeSH
• Saccharomyces cerevisiae účinky léků   MeSH
• Saccharomycetales chemie   metabolismus   MeSH
• Staphylococcus aureus účinky léků   MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• antiinfekční látky MeSH
• kultivační média MeSH

14-3-3 proteins bind phosphorylated binding partners to regulate several of their properties, including enzymatic activity, stability and subcellular localization. Here, two crystal structures are presented: the crystal structures of the 14-3-3 protein (also known as Bmh1) from the yeast Lachancea thermotolerans in the unliganded form and bound to a phosphopeptide derived from human PI4KB (phosphatidylinositol 4-kinase B). The structures demonstrate the high evolutionary conservation of ligand recognition by 14-3-3 proteins. The structural analysis suggests that ligand recognition by 14-3-3 proteins evolved very early in the evolution of eukaryotes and remained conserved, underlying the importance of 14-3-3 proteins in physiology.

• Klíčová slova
• 14-3-3 proteins, Bmh1, Bmh2, Lachancea thermotolerans, PI4KB, crystal structure, phosphopeptide,
• MeSH
• 1-fosfatidylinositol-4-kinasa chemie   genetika   metabolismus   MeSH
• Escherichia coli genetika   metabolismus   MeSH
• exprese genu MeSH
• fosfoproteiny chemie   genetika   metabolismus   MeSH
• fungální proteiny chemie   genetika   metabolismus   MeSH
• klonování DNA MeSH
• konformace proteinů, alfa-helix MeSH
• konzervovaná sekvence MeSH
• krystalografie rentgenová MeSH
• lidé MeSH
• ligandy MeSH
• molekulární evoluce MeSH
• molekulární modely MeSH
• plazmidy chemie   metabolismus   MeSH
• protein - isoformy chemie   genetika   metabolismus   MeSH
• proteiny 14-3-3 chemie   genetika   metabolismus   MeSH
• rekombinantní proteiny chemie   genetika   metabolismus   MeSH
• Saccharomycetales chemie   metabolismus   MeSH
• sekvence aminokyselin MeSH
• sekvenční seřazení MeSH
• strukturní homologie proteinů MeSH
• vazba proteinů MeSH
• vazebná místa MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• 1-fosfatidylinositol-4-kinasa MeSH
• fosfoproteiny MeSH
• fungální proteiny MeSH
• ligandy MeSH
• protein - isoformy MeSH
• proteiny 14-3-3 MeSH
• rekombinantní proteiny MeSH

Two hypotheses were tested: (1) microbial dephosphorylation of phytate in the presence of Ca²+ ions will result in the precipitation of hydroxyapatite-like crystals and (2) precipitation of calcium-phosphate crystals on and between sand-like particles can cause cementation. A growing culture of the dimorphic phytase-active yeast Arxula adeninivorans was introduced into a column filled with quartz particles and subsequently a liquid growth medium amended with calcium phytate was pumped through the column resulting in increased strength and stiffness of the quartz particle matrix. Environmental scanning electron microscope analysis combined with energy-dispersive X-ray measurement revealed cementation of the quartz particles by calcium-phosphate crystals. This microbial mineralization process could provide a novel approach to improving the mechanical properties like strength and stiffness of sandy soils.

• MeSH
• 6-fytasa metabolismus   MeSH
• fosforečnany vápenaté metabolismus   MeSH
• mikroskopie elektronová rastrovací MeSH
• půda analýza   MeSH
• půdní mikrobiologie * MeSH
• Saccharomycetales enzymologie   metabolismus   MeSH
• spektrometrie rentgenová emisní MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• 6-fytasa MeSH
• calcium phosphate MeSH Prohlížeč
• fosforečnany vápenaté MeSH
• půda MeSH

In this paper we report on searching for suitable reporters to monitor gene expression and protein secretion in the amylolytic yeast Schwanniomyces occidentalis. Several potential reporter and marker genes, formerly shown to be functional in other yeasts, were cloned downstream from the homologous invertase gene (INV) promoter and their activity was followed in conditions of repression and derepression of the INV promoter. However, neither beta-glucuronidase nor beta-lactamase nor phleomycin resistance-conferring gene, all originating from E. coli, were expressed in S. occidentalis cells to such a level to allow for monitoring of their activity. All the reporter genes tested have a higher percentage of GC (47-62%) in their DNA compared to the DNA composition of S. occidentalis genes that are more AT-rich (36% GC). The codon usage of all the reporter genes also varies from that of 16 so far sequenced S. occidentalis genes. This suggests that an appropriate composition of DNA and a codon usage similar to S. occidentalis genes might be very important parameters for an efficient expression of a heterologous gene in Schwanniomyces occidentalis. Indeed, two genes originating from Staphylococcus aureus, with an AT-content in their DNA similar to that of S. occidentalis, were functionally expressed in S. occidentalis cells. Both a phleomycin resistance-conferring gene and a chloramphenicol acetyltransferase-encoding gene thus represent suitable reporters of gene expression and protein secretion in S. occidentalis. Additionally, we show in this work that the transcription-regulating region and the signal peptide sequence of the S. occidentalis invertase gene were efficient to direct gene expression and subsequent protein secretion in Saccharomyces cerevisiae.

• MeSH
• bakteriální proteiny genetika   metabolismus   MeSH
• beta-laktamasy genetika   metabolismus   MeSH
• chloramfenikol-O-acetyltransferasa genetika   metabolismus   MeSH
• DNA fungální chemie   genetika   MeSH
• glukuronidasa genetika   metabolismus   MeSH
• glykosidhydrolasy genetika   metabolismus   MeSH
• invertasa MeSH
• kodon chemie   genetika   MeSH
• northern blotting MeSH
• promotorové oblasti (genetika) genetika   MeSH
• regulace genové exprese u hub MeSH
• reportérové geny genetika   MeSH
• Saccharomycetales genetika   metabolismus   MeSH
• zastoupení bazí MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• bakteriální proteiny MeSH
• beta-laktamasy MeSH
• chloramfenikol-O-acetyltransferasa MeSH
• DNA fungální MeSH
• glukuronidasa MeSH
• glykosidhydrolasy MeSH
• invertasa MeSH
• kodon MeSH
• phleomycin resistance protein, Streptoalloteichus hindustanus MeSH Prohlížeč

Distribution of microtubules and F-actin in aerobically growing cells of Dipodascus magnusii, belonging to the class Saccharomycetes was analyzed using immunofluorescence microscopy and labeling with rhodamine-tagged phalloidin. A conspicuous system of permanent cytoplasmic microtubules was observed in association with multiple nuclei. In elongating cells, helices of cytoplasmic microtubules appeared at the cell cortex. In cells approaching cytokinesis transversely oriented microtubules were revealed at incipient division sites. Confocal laser scanning microscopy showed a continuity of these transverse microtubules with the remaining microtubule network. The actin system of D. magnusii consisted of patches and filaments. Patches were found to accumulate at the tips of growing cells. Bands of fine actin filaments were usually observed before F-actin rings were established. A close cortical association of microtubules with the F-actin ring was documented on individual optical sections of labeled cells. Cells with developing septa showed medial F-actin discs associated at both sides with microtubules. Colocalization of cytoplasmic microtubules with actin filaments at the cortex of dividing cells supports a role of both cytoskeletal components in controlling cell wall growth and septum formation in D. magnusii.

• MeSH
• aktiny metabolismus   MeSH
• buněčné dělení MeSH
• faloidin MeSH
• fluorescenční protilátková technika MeSH
• konfokální mikroskopie MeSH
• mikrotubuly ultrastruktura   MeSH
• rhodaminy MeSH
• Saccharomycetales metabolismus   ultrastruktura   MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• aktiny MeSH
• faloidin MeSH
• rhodaminy MeSH

In the facultatively anaerobic yeast Saccharomyces cerevisiae the uptake rate and the accumulation ratio of 2-aminoisobutyric acid was decreased by some 30% by Fenton's reagent (FR), a powerful source of OH. radicals. Likewise, the uptake of glutamic acid, leucine and arginine was diminished. The mediated diffusion of 6-deoxy-D-glucose was not affected. The H+ symport of maltose and trehalose was inhibited by some 40% both in the initial rate and in the accumulation ratio. FR had a dramatic inhibitory effect when present during preincubation with 50 mmol/L glucose. In the obligately aerobic Lodderomyces elongisporus the uptake of all amino acids tested was decreased by 15-30%, that of 6-deoxy-D-glucose by about 10%. The initial rates of uptake of maltose and trehalose were depressed by FR by 40% and the acceleration of uptake observed after 8 min of incubation, was abolished by FR completely. Acidification rate of the external medium by S. cerevisiae in the presence of glucose or galactose was enhanced three-fold, that after subsequently added K+ was substantially decreased. FR appears to have a dual effect on sugar and amino acid transport processes in yeast: (1) it blocks carrier protein synthesis; (2) it inhibits the source of energy for transport. It does not appreciably affect the carrier proteins themselves.

• MeSH
• aktivní transport účinky léků   MeSH
• aminokyseliny metabolismus   MeSH
• fungální proteiny metabolismus   MeSH
• kinetika MeSH
• koncentrace vodíkových iontů MeSH
• metabolismus sacharidů MeSH
• peroxid vodíku farmakologie   MeSH
• Saccharomyces cerevisiae účinky léků   metabolismus   MeSH
• Saccharomycetales účinky léků   metabolismus   MeSH
• transportní proteiny metabolismus   MeSH
• železo farmakologie   MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• aminokyseliny MeSH
• Fenton's reagent MeSH Prohlížeč
• fungální proteiny MeSH
• peroxid vodíku MeSH
• transportní proteiny MeSH
• železo MeSH