Nadezhda V. Psurtseva

New efficient producers of fungal laccases

Two promising strains of laccase producers—Lentinus strigosus 1566 and Steccherinum ochraceum 1833—were found by screening of basidiomycetes. The cultivation conditions increasing the enzyme yield were selected. The maximal laccase activity was observed in the case of submerged cultivation of the mycelium immobilized on polycaproamide fibers in rich media in the presence of 2 mM CuSO4 in combination with the optimal inducer, namely, 2,6-dimethylphenol for L. strigosus and 2,4-dimethylphenol for S. ochraceum. Under these conditions, the activity of S. ochraceum laccase amounted to 33.1 U/ml and that of L. strigosus, to 186.5 U/ml. Anthracene was transformed with S. ochraceum laccase, and its oxidation to anthraquinone was demonstrated by mass spectrometry.

Comparative Analysis of the Ligninolytic Potential of Basidiomycetes Belonging to Different Taxonomic and Ecological Groups

Screening of the ligninolytic activity of basidiomycetes from the Komarov Botanical Institute Basidiomycetes Culture Collection (LE-BIN), Russian Academy of Sciences, belonging to different taxonomic and ecological groups was performed. The patterns of the position of taxa of active producers of ligninolytic enzymes in the modern system of fungi were identified. Cluster analysis showed that the group of fungi with the greatest ligninolytic and degradation potential includes representatives of the families Pleurotaceae, Polyporaceae, and Phanerochaetaceae, which perform the first stages of wood decomposition. As a result, species of interest for the further study of their oxidative potential and use in biotechnology were selected.

Diacylglyceryltrimethylhomoserine content and gene expression changes triggered by phosphate deprivation in the mycelium of the basidiomycete Flammulina velutipes

Diacylglyceryltrimethylhomoserines (DGTS) are betaine-type lipids that are phosphate-free analogs of phosphatidylcholines (PC). DGTS are abundant in some bacteria, algae, primitive vascular plants and fungi. In this study, we report inorganic phosphate (Pi) deficiency-induced DGTS synthesis in the basidial fungus Flammulina velutipes (Curt.: Fr.) Sing. We present results of an expression analysis of the BTA1 gene that codes for betaine lipid synthase and two genes of PC biosynthesis (CHO2 and CPT1) during phosphate starvation of F. velutipes culture. We demonstrate that FvBTA1 gene has increased transcript abundance under phosphate starvation. Despite depletion in PC, both CHO2 and CPT1 were determined to have increased expression. We also describe the deduced amino acid sequence and genomic structure of the BTA1 gene in F. velutipes. Phylogenetic relationships between putative orthologs of BTA1 proteins of basidiomycete fungi are discussed.

Draft Genome Sequence of the Fungus Trametes hirsuta 072

ABSTRACT A standard draft genome sequence of the white rot saprotrophic fungus Trametes hirsuta 072 (Basidiomycota, Polyporales) is presented. The genome sequence contains about 33.6 Mb assembled in 141 scaffolds with a G+C content of ~57.6%. The draft genome annotation predicts 14,598 putative protein-coding open reading frames (ORFs).

Alterations in the composition of membrane glycero-and sphingolipids in the course of Flammulina velutipes surface culture development.

Qualitative and quantitative characteristics of the alterations in the lipid composition of the membrane of the basidial fungus Flammulina velutipes in the course of surface culture development were investigated. Modifications of the lipid composition were shown to be timed to specific ontogeny stages, such as changes in the growth rate of the colonies, the appearance of differentiated vegetative cells, and the formation of generative structures. A slowdown of growth correlated with an alteration in the ratio of major classes of phospholipids, namely, with a decrease of phosphatidylcholine relative content and an increase in phosphatidylethanolamines. The differentiation of vegetative cells of the mycelium proceeded along with modifications of molecular composition of glycoceramides. In the course of the first week of growth, the surface culture of F. velutipes produced monohexosylceramides with epoxidized methyl sphingadienine as a sphingoid base. Later on, along with culture growth and specialization of mycelium cells, molecular species with methyl sphingadienine, common for basidiomycetes, start to prevail among the fungal glycoceramides. The formation of fruit bodies is accompanied by enrichment of molecules of phospholipids, mainly, the phosphatidylcholines, with unsaturated fatty acids.

Effect of submerged cultivation conditions and inducers on biosynthesis of extracellular laccase by a Trametes versicolor 1666 strain

Genetic analysis of basidiomycete Trametes versicolor 1666 from the Komarov Botanical Institute (LEBIN) Collection has been carried out that verified the phylogenetic position of the strain and revealed the presence of a single laccase gene. A study of cultivation dynamics of basidiomycete showed a low level of laccase, peroxidase, and Mn-peroxidase production on glucose-peptone and mineral media. The level of laccase activity in the culture liquid (CL) remained practically unchanged during cultivation in a medium optimized using full factorial experiment as compared to the standard medium. An addition of laccase inducers (syringaldazine, caffeic acid, and synaptic acid) had no effect on the enzyme activity in the culture liquid. Real time PCR showed a lack of reliable difference in the level of laccase gene expression when cultivating the strain under optimal conditions with and without copper ions in the medium. The transcription behavior of laccase gene and occurrence of enzyme activity when cultivating T. versicolor 1666 strain testify to the enzyme constitutive synthesis and the existence of additional regulatory ways in the laccase gene expression.

Formation of Diacylglyceryltrimethylhomoserines in the Surface Culture of the Basidiomycete Flammulina velutipes

Betaine-type lipids—diacylglyceryltrimethylhomoserines (DGTS)—were revealed in the mycelium of the basidial fungus Flammulina velutipes obtained by surface cultivation on agarized malt extract. DGTS accumulation was shown to occur at the late stages of culture development under deficiency of a complex of nutrients, including nitrogen, phosphorus, potassium, and trace elements. Induction of the synthesis of betaine lipids in F. velutipes occurred against the background of a decreased rate of growth of the vegetative mycelium, formation of monilioid hyphae, and inhibition of fructification. The relationship between DGTS formation and the environmental factors (temperature, illumination) was studied. It was established that the most active DGTS accumulation occurred at 15°C in the dark.

White-rot basidiomycetes Junghuhnia nitida and Steccherinum bourdotii: Oxidative potential and laccase properties in comparison with Trametes hirsuta and Coriolopsis caperata

White-rot basidiomycetes from the poorly studied residual polyporoid clade of Polyporales order Junghuhnia nitida (Pers.) Ryvarden and Steccherinum bourdotii Saliba & A. David grow as secondary xylotrohps on well decomposed woody materials. The main objective of the current study was to compare oxidative potential, growth, production of oxidative enzymes and laccase properties of J. nitida and S. bourdotii with that of typical primary xylotrohps Trametes hirsuta (Wulfen) Lloyd and Coriolopsis caperata (Berk.) Murrill, belonging to the core polyporoid clade. For the first time we report species J. nitida and S. bourdotii as active laccase producers. New laccases from J. nitida and S. bourdotii were purified and characterized. They had an identical molecular weight of 63 kDa and isoelectric points of 3.4 and 3.1, respectively. However, the redox potential of the T1 copper site for both J. nitida (610 mV) and S. bourdotii (640 mV) laccases was lower than those for T. hirsuta and C. caperata laccases. The new laccases showed higher temperature optima and better thermal stability than T. hirsuta and C. caperata laccases. Their half-lives were more than 40 min at 70 °C. The laccases from J. nitida and S. bourdotii showed higher affinity to syringyl-type phenolic compounds than T. hirsuta and C. caperata laccases. The oxidative potential of studied fungi as well as the properties of their laccases are discussed in terms of the fungal life-style.

Typification and Characterization of Trametes multicolor (Agaricomycetes), a Perspective Species of Medicinal Mushrooms

Nomenclature revision and enlarged taxonomical descriptions are still needed for some well-known species whose interpretation is complicated by many nomenclature or taxonomical problems. The polyporoid fungus widely known as Trametes ochracea (= Coriolus zonatus) belongs to such a problematic group. At the same time, recent data show that this species, like its sister species T. versicolor, seems to be a perspective subject for fungal biotechnology and pharmacology. This article is devoted to stabilizing the nomenclature of the species in question via lectotypification and epitypification of Boletus multicolor. It will clarify the name T. multicolor as applied to this species is nomenclaturally correct and useful, free of further problems. An expanded species description and cultural characterization of epitype materials are presented.