Advances in the Regulation and Production of Fungal Enzymes by Transcriptomics, Proteomics and Recombinant Strains Design

The goal of the review is description of the main characteristics of creatinine deiminase (CDI), an important bioanalytical tool for creatinine (Crn) assay. Crn is an essential metabolite for diagnostics of kidney disfunction and some other diseases, a biomarker to control the hemodialysis procedure, as well as an important analyte for sport medicine (estimation of general physiological status of athletes). We have described the important sources for CDI isolation, cloning of the corresponding gene, the construction of microbial recombinant strains, overproducing CDI, and characteristics of the enzyme from different microorganisms. There are reviewing also the new bioanalytical methods for quantitative determination of Crn, including enzymatic ones based on using CDI.

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MEIOTIC-LIKE RECOMBINATION IN VEGETATIVE DIKARYONS OF SCHIZOPHYLLUM COMMUNE

Genetics ◽

10.1093/genetics/92.4.1121 ◽

1979 ◽

Vol 92 (4) ◽

pp. 1121-1126

Author(s):

Carl Frankel

Keyword(s):

Schizophyllum Commune ◽

Genetic Selection ◽

Recombinant Strains

ABSTRACT A total of 65 Schizophyllum commune dikaryons of three different geno-types were synthesized, maintained on agar plates, prevented from fruiting, and macerated. The macerate was plated. Germlings of homokaryotic morphology were isolated using no chemical or genetic selection, grown, and tested for recombinant genotypes. Crossover types were plentiful among the homokaryons with recombinant genotypes, indicating that meiotic-like processes in vegetative dikaryons are common and may be as important as conventional basidial meiosis in the origin of recombinant strains.

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Arcopilus aureus MaC7A as a New Source of Resveratrol: Assessment of Amino Acid Precursors, Volatiles, and Fungal Enzymes for Boosting Resveratrol Production in Batch Cultures

Applied Sciences ◽

10.3390/app11104583 ◽

2021 ◽

Vol 11 (10) ◽

pp. 4583

Author(s):

Nemesio Villa-Ruano ◽

Luis Ángel Morales-Mora ◽

Jenaro Leocadio Varela-Caselis ◽

Antonio Rivera ◽

María de los Ángeles Valencia de Ita ◽

...

Keyword(s):

Amino Acid ◽

Filamentous Fungi ◽

Hydrolytic Enzymes ◽

Glucanase Activity ◽

Fungal Enzymes ◽

Activity Increase ◽

Batch Cultures ◽

Low Concentrations ◽

Chemical Factors ◽

Amino Acid Precursors

The chemical factors that regulate the synthesis of resveratrol (RV) in filamentous fungi are still unknown. This work reports on the RV production by Arcopilus aureus MaC7A under controlled conditions and the effect of amino acid precursors (PHE and TYR), monoterpenes (limonone, camphor, citral, thymol, menthol), and mixtures of hydrolytic enzymes (Glucanex) as elicitors for boosting fungal RV. Batch cultures with variable concentrations of PHE and TYR (50–500 mg L−1) stimulated RV production from 127.9 ± 4.6 to 221.8 ± 5.2 mg L−1 in basic cultures developed in PDB (pH 7) added with 10 g L−1 peptone at 30 °C. Maximum levels of RV and biomass were maintained during days 6–8 under these conditions, whereas a dramatic RV decrease was observed from days 10–12 without any loss of biomass. Among the tested volatiles, citral (50 mg L−1) enhanced RV production until 187.8 ± 2.2 mg L−1 in basic cultures, but better results were obtained with Glucanex (100 mg L−1; 198.3 ± 7.6 mg L−1 RV). Optimized batch cultures containing TYR (200 mg L−1), citral (50 mg L−1), thymol (50 mg L−1), and Glucanex (100 mg L−1) produced up to 237.6 ± 4.7 mg L−1 of RV. Our results suggest that low concentrations of volatiles and mixtures of isoenzymes with β-1, 3 glucanase activity increase the biosynthesis of fungal RV produced by A. aureus MaC7A in batch cultures.

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Correlation Study Between Biochemical and Molecular Pathways of Trichoderma harzianum Recombinant Strains on Plant Growth and Health

Journal of Plant Growth Regulation ◽

10.1007/s00344-021-10396-1 ◽

2021 ◽

Author(s):

Negin Eslahi ◽

Mojegan Kowsari ◽

Mohammad Reza Zamani ◽

Mostafa Motallebi

Keyword(s):

Plant Growth ◽

Trichoderma Harzianum ◽

Correlation Study ◽

Molecular Pathways ◽

Recombinant Strains

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The Parasexual Cycle in Candida albicans Provides an Alternative Pathway to Meiosis for the Formation of Recombinant Strains

PLoS Biology ◽

10.1371/journal.pbio.0060110 ◽

2008 ◽

Vol 6 (5) ◽

pp. e110 ◽

Cited By ~ 217

Author(s):

Anja Forche ◽

Kevin Alby ◽

Dana Schaefer ◽

Alexander D Johnson ◽

Judith Berman ◽

...

Keyword(s):

Candida Albicans ◽

Alternative Pathway ◽

Parasexual Cycle ◽

Recombinant Strains

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Construction and Functional Analysis of Mycolicibacterium smegmatis Recombinant Strains Carrying the Genes of Bacillary Cytochromes CYP106A1 and CYP106A2

Biotekhnologiya ◽

10.21519/0234-2758-2021-37-6-34-47 ◽

2021 ◽

Vol 37 (6) ◽

pp. 34-47

Author(s):

V.M. Nikolaeva ◽

V.V. Fokina ◽

A.A. Shutov ◽

A.V. Kazantsev ◽

N.I. Strizhov ◽

...

Keyword(s):

Cytochrome P450 ◽

Heterologous Expression ◽

Bacillus Megaterium ◽

Cytochromes P450 ◽

Maximum Level ◽

13C Nmr Spectroscopy ◽

Cytochrome P450 Monooxygenases ◽

Russian Science ◽

Steroid Nucleus ◽

Recombinant Strains

Mycolicibacterium smegmatis mc2155 has been genetically modified to be used as a platform for the expression of foreign cytochrome P450 monooxygenases by introducing deletions in the kshB and kstD genes that encode key stages of the enzymatic destruction of the steroid nucleus. Three sets of genetic constructs have been created for heterologous expression of the genes of cytochromes P450 CYP106A1 from Bacillus megaterium DSM319 and CYP106A2 from Bacillus megaterium ATCC13368 in Mycolicibacterium smegmatis mc2155 (ΔkshBΔkstD) cells. The recombinant plasmids contained monocistronic expression cassettes of cytochrome genes (NS31 and pNS32), or tricistronic cassettes of cytochrome genes together with cDNA copies of adrenodoxin and andrenodoxin reductase genes of the bovine adrenal cortex (pNS33 and pNS34), or monocistronic gene cassettes of chimeric cytochromes fused with the DNA sequence encoding the CYP116B2 reductase domain from the soil bacterium Rhodococcus sp. NCIMB 9784 (pNS35 and pNS36). The recombinant strains of mycolicibacteria were shown to selectively monohydroxylate androstenedione (AD) under growth conditions. The product was identified as 15-hydroxyandrostenedione (15-OH-AD) by mass spectrometry and 1H and 13C NMR spectroscopy. The maximum level of 15-OH-AD production (17.3 ± 1.5 mg/L) was observed when using the recombinant M. smegmatis mc2155 (ΔkshBΔkstD) (pNS32) strain, which expresses a single cyp106A2 gene from B. megaterium ATCC13368. Host proteins of M. smegmatis mc2155 were shown to be capable of supplying electrons to heterologous cytochromes to support their hydroxylating activity. The results are of priority character, expand the understanding of the hydroxylation of steroid compounds by bacterial cytochromes CYP106A1/A2 and are important for the creation of microbial strains producing valuable hydroxysteroids. cyp106A1, cyp106A2, cytochrome P450, heterologous expression, Bacillus megaterium, Mycolicibacterium smegmatis, 15β-hydroxylation, bioconversion, steroids This work was supported by the Russian Science Foundation (project No. 21-64-00024).

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Amylase hyper-producing haploid recombinant strains of Thermomyces lanuginosus obtained by intraspecific protoplast fusion

Canadian Journal of Microbiology ◽

10.1139/w00-042 ◽

2000 ◽

Vol 46 (7) ◽

pp. 669-673 ◽

Cited By ~ 3

Author(s):

K Rubinder ◽

B S Chadha ◽

S Singh ◽

H S Saini

Keyword(s):

Protoplast Fusion ◽

Culture Filtrate ◽

Catabolite Repression ◽

Type Strain ◽

Wild Type Strain ◽

Thermomyces Lanuginosus ◽

Wild Type ◽

Haploid Strain ◽

Recombinant Strains ◽

Specific Activities

Amylase hyper-producing, catabolite-repression-resistant, recombinant strains were produced by intraspecific protoplast fusion of thermophilic fungus Thermomyces lanuginosus strains, using well-characterized, morphological, and 2-deoxy-D-glucose resistant markers. The fusant heterokaryons exhibited enhanced amylase activities as compared to the amylase hyper-producing parental strain (T2). Diploids derived from heterokaryons segregated to stable haploid recombinant strains. In the haploid strain (Tlh 4q), approximately 5-fold higher specific activities of α-amylase and glucoamylase in the culture filtrate were observed as compared to the wild-type strain (W0).Key words: Thermomyces lanuginosus, protoplast fusion, amylase hyper-producing strain, catabolite repression.

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