Characterization of the $\alpha$-mannosidase Gene Family in Filamentous Fungi: N-glycan Remodelling for the Development of Eukaryotic Expression Systems

Eades, C.Joshua,Hintz, William E. The Korean Society for Biotechnology and Bioengine 2000 Biotechnology and Bioprocess Engineering Vol.5 No.4

Although filamentous fungi are used extensively for protein expression, their use for the production of heterologous glycoproteins is constrained by the types of N-glycan structures produced by filamentous fungi as compared to those naturally found on the glycoproteins. Attempts are underway to engineer the N-glycan synthetic pathways in filamentous fungi in order to produce fungal expression strains which can produce heterologous glycoproteins carrying specific N-glycan structures. To fully realize this goal, a detailed understanding of the genetic components of this pathway in filamentous fungi is required. In this review, we discuss the characterization of the $\alpha$-mannosidase gene family in filamentous fungi and its implications for the elucidation of the N-glycan synthetic pathway.

Morphological Engineering of Filamentous Fungi: Research Progress and Perspectives

Zhengwu Lu,Zhiqun Chen,Yunguo Liu,Xuexue Hua,Cuijuan Gao,Jingjing Liu The Korean Society for Microbiology and Biotechnol 2024 Journal of microbiology and biotechnology Vol.34 No.6

Filamentous fungi are important cell factories for the production of high-value enzymes and chemicals for the food, chemical, and pharmaceutical industries. Under submerged fermentation, filamentous fungi exhibit diverse fungal morphologies that are influenced by environmental factors, which in turn affect the rheological properties and mass transfer of the fermentation system, and ultimately the synthesis of products. In this review, we first summarize the mechanisms of mycelial morphogenesis and then provide an overview of current developments in methods and strategies for morphological regulation, including physicochemical and metabolic engineering approaches. We also anticipate that rapid developments in synthetic biology and genetic manipulation tools will accelerate morphological engineering in the future.

Lipase-producing Filamentous Fungi from Non-dairy Creamer Industrial Waste

Triyaswati, Desty,Ilmi, Miftahul The Korean Society for Microbiology and Biotechnol 2020 한국미생물·생명공학회지 Vol.48 No.2

Lipase-producing fungi have been isolated from environments containing lipids. The non-dairy creamer industrial waste has a high amount of lipids so it is a potential source for the isolation of lipase-producing fungi. However, the study of fungi that secrete lipase from this industrial waste has not been reported. The purpose of this study was to obtain lipase-producing filamentous fungi from non-dairy creamer industrial waste. Mineral salt and potato dextrose agar were used as media for the isolation process. The qualitative screening was conducted using phenol red agar medium and the quantitative screening using broth medium containing glucose and olive oil. Isolates producing the highest amounts of lipase were identified with molecular methods. We found that 5 out of 19 isolated filamentous fungi are lipase producers. Further analysis showed that isolate Ms.11 produced the highest amount of lipase compared to others. Based on ITS sequence Ms.11 was identified as Aspergillus aculeatus. The lipase activity in medium containing 1% glucose + 1% olive oil at pH 7.0 and 30℃ after 96 and 120 h of incubation was 5.13 ± 0.30 U/ml and 5.22 ± 0.59 U/ml, respectively. The optimum lipase activity was found at pH 7.0, 30℃ and using methanol or ethanol in the reaction tube. Lipase was more stable at 20-30℃ and maintained 85% of its activity. It was concluded that isolate Ms.11 is a potential source of lipase that catalyzes transesterification reactions. Further studies are required to optimize lipase production to make the strain suitable for industry purposes.

Lipase-producing Filamentous Fungi from Non-dairy Creamer Industrial Waste

Desty Triyaswati,Miftahul Ilmi 한국미생물·생명공학회 2020 한국미생물·생명공학회지 Vol.48 No.2

Lipase-producing fungi have been isolated from environments containing lipids. The non-dairy creamer industrial waste has a high amount of lipids so it is a potential source for the isolation of lipase-producing fungi. However, the study of fungi that secrete lipase from this industrial waste has not been reported. The purpose of this study was to obtain lipase-producing filamentous fungi from non-dairy creamer industrial waste. Mineral salt and potato dextrose agar were used as media for the isolation process. The qualitative screening was conducted using phenol red agar medium and the quantitative screening using broth medium containing glucose and olive oil. Isolates producing the highest amounts of lipase were identified with molecular methods. We found that 5 out of 19 isolated filamentous fungi are lipase producers. Further analysis showed that isolate Ms.11 produced the highest amount of lipase compared to others. Based on ITS sequence Ms.11 was identified as Aspergillus aculeatus. The lipase activity in medium containing 1% glucose + 1% olive oil at pH 7.0 and 30℃ after 96 and 120 h of incubation was 5.13 ± 0.30 U/ml and 5.22 ± 0.59 U/ml, respectively. The optimum lipase activity was found at pH 7.0, 30℃ and using methanol or ethanol in the reaction tube. Lipase was more stable at 20−30℃ and maintained 85% of its activity. It was concluded that isolate Ms.11 is a potential source of lipase that catalyzes transesterification reactions. Further studies are required to optimize lipase production to make the strain suitable for industry purposes.

Alternaria brassicicola SW-3의 항진균 활성

정동선,라여정,변한나,정은영 서울여자대학교 자연과학연구소 2003 자연과학연구논문집 Vol.15 No.-

The fungus Alternaria brassicicola is a causal agent of black leaf spot in. several cruciferous plants. We found A. brassicicola SW-3 produced antifungal metabolite, in addition to depudecin, reported as a histone deacetylase inhibitor. The metabolite produced by A. brassicicola SW-3 exhibited antifungal activity against various filamentous fungi such as Rhizopus, Mucor, Aspergillus, Microsporium, and Trichophyton. The antifungal metabolite was extracted with ethyl acetate from the culture filtrate of A. brassicicola, and purified through silica gel column chromatography. The purified compound exhibited a broad antifungal spectrum, particularly potent activity against Microsporum canis and Trichophyton rubrum which are dermatophytes associated with cutaneous Mycosiscause. The purified metabolite showed very complex spectral data from proton and carbon-13 NMR analysis, indicating the compound contains both lipid and carbohydrate moiety. Although structure of the antifungal metabolite is not elucidated yet, these preliminary results suggest that the antifungal metabolite from A. brassicicola could be used as a lead compound for treatment of skin diseases caused by filamentous fungi.

Mini-review : Microcyle Conidiation in Filamentous Fungi

( Bok Nam Jung ),( So Yeon Kim ),( Jung Kwan Lee ) 한국균학회 2014 Mycobiology Vol.42 No.1

The typical life cycle of filamentous fungi commonly involves asexual sporulation after vegetative growth in response to environmental factors.The production of asexual spores is critical in the life cycle of most filamentous fungi.Normally, conidia are produced from vegetative hyphae(termed mycelia).However, fungal species subjected to stress conditions exhibit an extremely simplified asexual life cycle, in which the conidia that germinate directly generate further conidia, without forming mycelia.This phenomenon has been termed as microcycle conidiation, and to date has been reported in more than 100 fungal species.In this review, first, we present the morphological properties of fungi during microcycle conidiation, and dividemicrocycle conidiation into four simple categories, even though fungal species exhibit a wide variety of morphological differences during micro cycle conidiogenesis. Second, we describe the factors that influence microcycle conidiation in various fungal species, and present recent genetic studies that have identified the genes responsible for this process.Finally, we discuss the biological meaning and application of micro cycle conidiation.

Study of the Rheological Properties of a Fermentation Broth of the Fungus Beauveria bassiana in a Bioreactor Under Different Hydrodynamic Conditions

Nunez-Ramirez, Diola Marina,Medina-Torres, Luis,Valencia-Lopez, Jose Javier,Calderas, Fausto,Lopez-Miranda, Javier,Medrano-Roldan, Hiram,Solis-Soto, Aquiles The Korean Society for Microbiology and Biotechnol 2012 Journal of microbiology and biotechnology Vol.22 No.11

Fermentation with filamentous fungi in a bioreactor is a complex dynamic process that is affected by flow conditions and the evolution of the rheological properties of the medium. These properties are mainly affected by the biomass concentration and the morphology of the fungus. In this work, the rheological properties of a fermentation with the fungus Beauveria bassiana under different hydrodynamic conditions were studied and the rheological behavior of this broth was simulated through a mixture of carboxymethyl cellulose sodium and cellulose fibers (CMCNa-SF). The bioreactor was a 10 L CSTR tank operated at different stir velocities. Rheological results were similar at 100 and 300 rpm for both systems. However, there was a significant increase in the viscosity accompanied by a change in the consistence index, calculated according to the power law model, for both systems at 800 rpm. The systems exhibited shear-thinning behavior at all stir velocities, which was determined with the power law model. The mixing time was observed to increase as the cellulose content in the system increased and, consequently, the efficiency of mixing diminished. These results are thought to be due to the rheological and morphological similarities of the two fungal systems. These results will help in the optimization of scale-up production of these fungi.

A CRISPR/Cas9 Cleavage System for Capturing Fungal Secondary Metabolite Gene Clusters

( Xinran Xu ),( Jin Feng ),( Peng Zhang ),( Jie Fan ),( Wen-bing Yin ) 한국미생물생명공학회(구 한국산업미생물학회) 2021 Journal of microbiology and biotechnology Vol.31 No.1

More and more available fungal genome sequence data reveal a large amount of secondary metabolite (SM) biosynthetic ‘dark matter’ to be discovered. Heterogeneous expression is one of the most effective approaches to exploit these novel natural products, but it is limited by having to clone entire biosynthetic gene clusters (BGCs) without errors. So far, few effective technologies have been developed to manipulate the specific large DNA fragments in filamentous fungi. Here, we developed a fungal BGC-capturing system based on CRISPR/Cas9 cleavage in vitro. In our system, Cas9 protein was purified and CRISPR guide sequences in combination with in vivo yeast assembly were rationally designed. Using targeted cleavages of plasmid DNAs with linear (8.5 kb) or circular (8.5 kb and 28 kb) states, we were able to cleave the plasmids precisely, demonstrating the high efficiency of this system. Furthermore, we successfully captured the entire Nrc gene cluster from the genomic DNA of Neosartorya fischeri. Our results provide an easy and efficient approach to manipulate fungal genomic DNA based on the in vitro application of Cas9 endonuclease. Our methodology will lay a foundation for capturing entire groups of BGCs in filamentous fungi and accelerate fungal SMs mining.

Global Approaches to Identify Genes Involved during Infection Structure Formation in Rice Blast Fungus,Magnaporthe grisea

최우봉 한국식물병리학회 2003 Plant Pathology Journal Vol.19 No.1

The ascomycete Magnaporthe grisea is a pathogen of rice blast and is known to form specialized infection structures called appressoria for successful infection into host cells. To understand the molecular mechanism underlying infection process, appressorium-related genes were identified through global approaches including EST sequencing, differential hybridization, and suppression subtractive hybridization. EST database was generated on >2,000 cDNA clones randomly selected from appressorium stage cDNA library. Large number of ESTs showed homology to known proteins possibly involved in infection-related cellular development (attachment, germination, appressorium formation, and colonization) of rice blast fungus. The 1051 ESTs showing significant homology to known genes were assigned to 11 functional categories. Differential hybridization and suppression subtractive hybridization were applied to identify genes showing an appressorium stage specific expression pattern. A number of genes were selected as up-regulated during appressorium formation compared with the vegetative growing stage. Clones from various cDNA libraries constructed in different developmental stages were arrayed on slide glass for further expression profiling study. Functional characterization of genes identified from these global approaches may lead to a better understanding of the infection process of this devastating plant disease, and the development of novel ways to protect host plant.

Research Notes : Filamentous Fungi Isolated from Platypus koryoensis, the Insect Vector of Oak Wilt Disease in Korea

( Dong Yeon Suh ),( Min Woo Hyun ),( Seong Hwan Kim ),( Sang Tae Seo ),( Kyung Hee Kim ) 한국균학회 2011 Mycobiology Vol.39 No.4

The ambrosia beetle, Platypus koryoensis, is a serious pest of oak trees in Korea. In this study we investigated filamentous fungi present in the body of the beetle. Fourteen genera of filamentous fungi belonging to Ascomycota and Basidiomycota were isolated. Among the isolated fungi, some were able to produce wood degrading enzymes. This is first report of fungi associated with P. koryoensis.