Journal of Microbiology

Research Article

Simultaneous gene editing of both nuclei in a dikaryotic strain of Ganoderma lucidum using Cas9-gRNA ribonucleoprotein: Yeon-Jae Choi, Hyerang Eom, Rutuja Nandre, Minseek Kim, Youn-Lee Oh, Sinil Kim, Hyeon-Su Ro; J. Microbiol. 2025;63(1):e.2409006. Published online January 24, 2025; DOI: https://doi.org/10.71150/jm.2409006

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Abstract PDF Supplementary Material: The presence of multiple nuclei in a common cytoplasm poses a significant challenge to genetic modification in mushrooms. Here, we demonstrate successful gene editing in both nuclei of a dikaryotic strain of Ganoderma lucidum using the Cas9-gRNA ribonucleoprotein complex (RNP). The RNP targeting the pyrG gene was introduced into dikaryotic protoplasts of G. lucidum, resulting in the isolation of 31 mycelial colonies resistant to 5-fluoroorotic acid (5-FOA). Twenty-six of these isolates were confirmed as dikaryotic strains by the presence of two distinct A mating type markers, denoted as A1 and A2. All dikaryons exhibited clamp connections on their mycelial hyphae, while the remaining 5 transformants were monokaryotic. Subsequent sequence analysis of PCR amplicons targeting pyrG revealed that two dikaryons harbored disrupted pyrG in both nuclei (pyrG-/pyrG-), while 10 and 14 displayed pyrG+/pyrG- (A1/A2) and pyrG-/pyrG+ (A1/A2) configurations, respectively. The disruption was achieved through non-homologous end joining repair, involving deletion or insertion of DNA fragments at the site of the double-strand break induced by RNP. Importantly, the nuclei were stable throughout 10 serial transfers over a period of 6 months. These findings highlight the capability of RNP to target genes across multiple nuclei within the same cytoplasm.

Journal Articles

Metabolomic profiling reveals enrichment of cordycepin in senescence process of Cordyceps militaris fruit bodies: Junsang Oh , Deok-Hyo Yoon , Bhushan Shrestha , Hyung-Kyoon Choi , Gi-Ho Sung; J. Microbiol. 2019;57(1):54-63. Published online December 29, 2018; DOI: https://doi.org/10.1007/s12275-019-8486-z

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Cordyceps militaris is a species of Cordyceps that is classified in the Cordycipitaceae family and is well known in East Asia as a traditional medicinal mushroom. Its artificial fruit body has been widely cultivated for commercial use in cosmetics, functional food, and medicine. To explore the metabolites associated with fruit body development, we conducted gas chromatography mass spectrometry (GC-MS) analyses based on developmental stage, which was divided into the growth period (stage 1, stage 2, and stage 3) and aging period (stage 4). We detected 39 biochemical metabolites associated with nucleotide, carbohydrate, and amino acid metabolism. Cordycepin, one of the representative bioactive compounds in C. militaris, was significantly enriched in stage 4 of aging period and is associated with glucose accumulation. The accumulation of cordycepin in stage 4 of aging period also seems to be related to the glutamine and glutamic acid pathway. Our
results
also showed enrichment of other bioactive compounds such as mannitol and xylitol in stage 4 of aging period. Our metabolomic profiling based on the developmental stages of C. militaris is useful for exploring bioactive compounds (e.g., cordycepin, mannitol, GABA, and xylitol) that are enriched in stage 4 of aging period and understanding the biosynthetic mechanisms associated with cordycepin production. Through optimization of fruit body cultivation by selecting stage 4 of aging period as a harvesting time, our findings can be utilized in food and medical applications of C. militaris in future.

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Core microbes in Cordyceps militaris sclerotia and their nitrogen metabolism-related ecological functions
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Frontiers in Microbiology.2024;[Epub] CrossRef
Current Progress Regarding Cordyceps militaris, Its Metabolite Function, and Its Production
Yu-Chieh Chou, Ting-Hsuan Sung, Shi-Jing Hou, Darin Khumsupan, Shella Permatasari Santoso, Kuan-Chen Cheng, Shin-Ping Lin
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Roypim Thananusak, Kobkul Laoteng, Nachon Raethong, Mattheos Koffas, Wanwipa Vongsangnak
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Functional genomics and systems biology of Cordyceps species for biotechnological applications
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Effect of Cultivation Stages of Hericium erinaceus on the Contents of Major Components and α-Glucosidase Inhibitory Activity
Se Hwan Ryu, Beom Seok Kim, Ayman Turk, Sang Won Yeon, Solip Lee, Hak Hyun Lee, Sung Min Ko, Bang Yeon Hwang, Kyeong Lee
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Weerasak Seelarat, Sujarinee Sangwanna, Teerapap Panklai, Natthaphon Chaosuan, Atipong Bootchanont, Chakkaphan Wattanawikkam, Ahkasit Subcharoen, Nuttapon Subcharoen, Nattapong Chanchula, Dheerawan Boonyawan, Porramain Porjai
Plasma Chemistry and Plasma Processing.2023; 43(1): 139. CrossRef
Cordycepin production by the potential fungal strains Cordyceps militaris BCC 2819 and Cordyceps cicadae BCC 19788 in submerged culture during batch and Fed-batch fermentation
Werapan Borworn, Nutaratat Pumin, Ariyaphuttarat Siriporn, Prathumpai Wai
African Journal of Biotechnology.2022; 21(10): 483. CrossRef
Research Progress on Cordycepin Synthesis and Methods for Enhancement of Cordycepin Production in Cordyceps militaris
Li Wang, Huanhuan Yan, Bin Zeng, Zhihong Hu
Bioengineering.2022; 9(2): 69. CrossRef
Application of Metabolomics in Fungal Research
Guangyao Li, Tongtong Jian, Xiaojin Liu, Qingtao Lv, Guoying Zhang, Jianya Ling
Molecules.2022; 27(21): 7365. CrossRef
Cordycepin production by a novel endophytic fungus Irpex lacteus CHG05 isolated from Cordyceps hawkesii Gray
Ziying Liu, Guihua Leng, Jingbai Wen, Ganqi Deng, Jiayao Jiang
Folia Microbiologica.2022; 67(6): 851. CrossRef
Heavy Metals, Halogenated Hydrocarbons, Phthalates, Glyphosate, Cordycepin, Alcohol, Drugs, and Herbs, Assessed for Liver Injury and Mechanistic Steps
Rolf Teschke, Tran Dang Xuan
Frontiers in Bioscience-Landmark.2022;[Epub] CrossRef
Effects on total phenolic and flavonoid content, antioxidant properties, and angiotensin I-converting enzyme inhibitory activity of beans by solid-state fermentation with Cordyceps militaris
Weiwei Liu, Mengqian Dun, Xinyuan Liu, Guoying Zhang, Jianya Ling
International Journal of Food Properties.2022; 25(1): 477. CrossRef
Effect of Spirulina biomass residue on yield and cordycepin and adenosine production of Cordyceps militaris culture
Siriluck Iamtham, Adisak Kaewkam, Sermsiri Chanprame, Wanida Pan-utai
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Phongsakorn Kunhorm, Phongsakorn Chueaphromsri, Nipha Chaicharoenaudomrung, Parinya Noisa
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Soo Kweon Lee, Ju Hun Lee, Hyeong Ryeol Kim, Youngsang Chun, Ja Hyun Lee, Hah Young Yoo, Chulhwan Park, Seung Wook Kim
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Root-associated bacteria influencing mycelial growth of Tricholoma matsutake (pine mushroom): Seung-Yoon Oh , Young Woon Lim; J. Microbiol. 2018;56(6):399-407. Published online June 1, 2018; DOI: https://doi.org/10.1007/s12275-018-7491-y

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Abstract

Tricholoma matsutake is an ectomycorrhizal fungus usually associated with Pinus densiflora in South Korea. Fruiting bodies (mushrooms) of T. matsutake are economically important due to their attractive aroma; yet, T. matsutake is uncultivatable and its habitat is rapidly being eradicated due to global climate change. Root-associated bacteria can influence the growth of ectomycorrhizal fungi that co-exist in the host rhizosphere and distinctive bacterial communities are associated with T. matsutake. In this study, we investigated how these bacterial communities affect T. matsutake growth by isolating bacteria from the roots of P. densiflora colonized by ectomycorrhizae of T. matsutake and co-culturing rootassociated bacteria with T. matsutake isolates. Thirteen species of bacteria (27 isolates) were found in pine roots, all belonging to the orders Bacillales or Burkholderiales. Two species in the genus Paenibacillus promoted the growth of T. matsutake in glucose poor conditions, likely using soluble metabolites. In contrast, other bacteria suppressed the growth of T. matsutake using both soluble and volatile metabolites. Antifungal activity was more frequent in glucose poor conditions. In general, pine rhizospheres harbored many bacteria that had a negative impact on T. matsutake growth and the few Paenibacillus species that promoted T. matsutake growth. Paenibacillus species, therefore, may represent a promising resource toward successful cultivation of T. matsutake.

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Research Support, Non-U.S. Gov'ts

Species identity of Phellinus linteus (sanghuang) extensively used as a medicinal mushroom in Korea: Jae-Gu Han , Min-Woo Hyun , Chang Sun Kim , Jong Won Jo , Jae-Han Cho , Kang-Hyo Lee , Won-Sik Kong , Sang-Kuk Han , Junsang Oh , Gi-Ho Sung; J. Microbiol. 2016;54(4):290-295. Published online April 1, 2016; DOI: https://doi.org/10.1007/s12275-016-5520-2

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Abstract

Sanghuang is a medicinal mushroom that has gained particular attention in Korea. It has been extensively studied for the past few decades as a natural immune booster and cancer suppressor. Although the scientific name, Phellinus linteus, has been commonly used to refer to the sanghuang mushroom, the species identity of sanghuang has been called into question due to the ambiguity of its circumscription and the inadequacy of morphological distinctions within allied species. Because the species concept of sanghuang has been elucidated by recent molecular phylogenetic studies, it has become necessary to clarify the taxonomic positions of sanghuang strains extensively utilized in Korea. We conducted a phylogenetic analysis of 74 strains belonging to the P. linteus-baumii complex based on ITS nrDNA sequences. Parental stains of sanghuang varieties formally registered in the Korea Seed & Variety Service, including ASI 26046 (Corea sanghuang), 26114 (Boolro), and 26115 (HK 1-ho) were grouped with Sanghuangporus sanghuang instead of P. linteus in the inferred phylogeny.

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Evaluation of antioxidant, anti-inflammatory and anti-proliferative activities of ethanol extracts from different varieties of Sanghuang species
Wang-Ching Lin, Jeng-Shyan Deng, Shyh-Shyun Huang, Sheng-Hua Wu, Hui-Yi Lin, Guan-Jhong Huang
RSC Advances.2017; 7(13): 7780. CrossRef

Pb Tolerance and Bioaccumulation by the Mycelia of Flammulina velutipes in Artificial Enrichment Medium: Changwei Zhu , Zhengpeng Li , Decai Li , Yan Xin; J. Microbiol. 2014;52(1):8-12. Published online January 4, 2014; DOI: https://doi.org/10.1007/s12275-014-2560-3

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Abstract

Mushrooms have the ability to accumulate high concentrations of heavy metals, which gives them potential for use as bioremediators of environmental contamination. The Pb2+ tolerance and accumulation ability of living mycelia of Flammulina velutipes were studied in this work. Mycelial growth was inhibited when exposed to 1 mM Pb2+. The colony diameter on solid medium decreased almost 10% compared with the control. Growth decreased almost 50% when the Pb2+ concentration increased to 4 mM in the medium, with the colony diameter decreasing from 80 mm to 43.4 mm, and dry biomass production in liquid cultures decreasing from 9.23±0.55 to 4.27±0.28 g/L. Lead ions were efficiently accumulated in the mycelia. The amount of Pb2+ in the mycelia increased with increasing Pb2+ concentration in the medium, with the maximum concentration up to 707±91.4 mg/kg dry weight. We also show evidence that a large amount of the Pb2+ was adsorbed to the mycelial surface, which may indicate that an exclusion mechanism is involved in Pb tolerance. These results demonstrate that F. velutipes could be useful as a remediator of heavy metal contamination because of the characteristics of high tolerance to Pb2+ and efficient accumulation of Pb2+ ions by the mycelia.

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An Approach to Evaluate Pb Tolerance and Its Removal Mechanisms by Pleurotus opuntiae
Priyanka Yadav, Vartika Mishra, Tejmani Kumar, Awadhesh Kumar Rai, Ayush Gaur, Mohan Prasad Singh
Journal of Fungi.2023; 9(4): 405. CrossRef
Detrimental and essential elements in fruiting bodies of wild-growing fungi Coprinus comatus, Flammulina velutipes, and Armillaria ostoyae
Jan Vondruška, Jan Šíma, Marek Kobera, Lukáš Rokos, Martin Šeda, Lubomír Svoboda
Journal of Environmental Science and Health, Part B.2022; 57(4): 243. CrossRef
Waste apple wood: A safe and economical alternative substrate for the cultivation of Pleurotus ostreatus and Lentinula edodes
Guorong Yu, Xinxin Li, Shengting Zhao, Shuguang Sun, Yanru Yu, Jinzhi Chen, Xianhao Cheng, Weihuan Li
Folia Horticulturae.2022; 34(2): 173. CrossRef
The effect of different substrates on the growth of six cultivated mushroom species and composition of macro and trace elements in their fruiting bodies
Marek Siwulski, Piotr Rzymski, Anna Budka, Pavel Kalač, Sylwia Budzyńska, Luiza Dawidowicz, Edmund Hajduk, Lidia Kozak, Joanna Budzulak, Krzysztof Sobieralski, Przemysław Niedzielski
European Food Research and Technology.2019; 245(2): 419. CrossRef
Biosorption characteristic of Alcaligenes sp. BAPb.1 for removal of lead(II) from aqueous solution
Yu Jin, Sumei Yu, Chunying Teng, Tao Song, Liying Dong, Jinsong Liang, Xin Bai, Xiuhong Xu, Juanjuan Qu
3 Biotech.2017;[Epub] CrossRef

Morphological and Genetic Characteristics of Newly Crossbred Cauliflower Mushroom (Sparassis latifolia): Hong-Duck Sou , Rhim Ryoo , Sung-Ryul Ryu , Kang-Hyeon Ka , Hyun Park , Sung-Hyun Joo; J. Microbiol. 2013;51(5):552-557. Published online June 25, 2013; DOI: https://doi.org/10.1007/s12275-013-2666-z

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Abstract

Cauliflower mushroom (Sparassis latifolia or S. crispa) is popular for food and medicine. Importance of new varieties of Sparassis was raised and studied widely by protection system of UPOV. In this study, 10 crossbred strains of Sparassis latifolia that specifically expressed distinctive features during basidiocarp formation and mycelium growth were applied to sawdust medium inoculated with S. latifolia mycelia. The 10 crossbred strains were divided into 3 groups on the basis of morphological (size of marginal wave and basidiocarp color) and genetic characteristics. Each phenotype of the parent and crossbred strains represented 3 marginal wave-sizes (large, medium, and small) and 3 color notations (NN155D, 163C, and 8D). Our result suggests that morphological characteristics of cauliflower mushroom can be affected by various environmental and genetic stimuli under artificial conditions such as crossbreed. Also this research showed genetic differences among breeding isolates and their morphological characteristics were correlated with the molecular data within parent and crossed strain.

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Neuroprotective Effects of Sparassis crispa Ethanol Extract through the AKT/NRF2 and ERK/CREB Pathway in Mouse Hippocampal Cells
Malk Eun Pak, Wei Li
Journal of Fungi.2023; 9(9): 910. CrossRef
Breeding of a high-yield strain for commercial cultivation by crossing Pholiota adiposa
Chengbo Rong, Shuang Song, Li Yang, Xuejiao Pan, Yu Liu, Shouxian Wang
Ciência e Agrotecnologia.2021;[Epub] CrossRef
Elicitor-induced β-glucan contents in fruit body of cauliflower mushroom (Sparassis latifolia)
Rhim Ryoo, Hong-Duck Sou, Kang-Hyeon Ka, Hyun Park
Forest Science and Technology.2018; 14(3): 119. CrossRef
Development of a highly productive strain of Pleurotus tuoliensis for commercial cultivation by crossbreeding
Shouxian Wang, Shuang Zhao, Zhenxing Huang, Limin Yin, Jie Hu, Jianghong Li, Yu Liu, Chengbo Rong
Scientia Horticulturae.2018; 234: 110. CrossRef
Effects of Sparassis crispa in Medical Therapeutics: A Systematic Review and Meta-Analysis of Randomized Controlled Trials
Le Thi Nhu Ngoc, You-Kwan Oh, Young-Jong Lee, Young-Chul Lee
International Journal of Molecular Sciences.2018; 19(5): 1487. CrossRef
The mycelial growth and ligninolytic enzyme activity of cauliflower mushroom (Sparassis latifolia)
Hong-Duck Sou, Rhim Ryoo, Kang-Hyeon Ka, Hyun Park
Forest Science and Technology.2017; 13(4): 158. CrossRef

Journal Article

Biochemical Characterization of Chitinase 2 Expressed during the Autolytic Phase of the Inky Cap, Coprinellus congregatus: Yuri Kang , Hyewon Kim , Hyoung T. Choi; J. Microbiol. 2013;51(2):189-193. Published online April 27, 2013; DOI: https://doi.org/10.1007/s12275-013-2535-9

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Abstract: Fungal cell walls consist of various glucans and chitin. The inky cap, Coprinellus congregatus, produces mushrooms at 25°C in a regime of 15 h light/9 h dark, and then the mushroom is autolyzed rapidly to generate black liquid droplets in which no cell walls are detected by microscopy. Chitinase cDNA from the mature mushroom tissues of C. congregatus, which consisted of 1,622 nucleotides (chi2), was successfully cloned using the rapid amplification of cDNA ends polymerase chain reaction technique. The deduced 498 amino acid sequence of Chi2 had a conserved catalytic domain as in other fungal chitinase family 18 enzymes. The Chi2 enzyme was purified from the Pichia pastoris expression system, and its estimated molecular weight was 68 kDa. The optimum pH and temperature of Chi2 was pH 4.0 and 35°C, respectively when 4-nitrophenyl N,N’-diacetyl-β-D-chitobioside was used as the substrate. The Km value and Vmax for the substrate A, 4-nitrophenyl N,N’-diacetyl-β-D-chitobioside, was 0.175 mM and 0.16 OD min-1unit-1, respectively.

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Isolation and Characterization of a Mycovirus in Lentinula edodes: Hyo-Kyoung Won , So-Jung Park , Dong-Kyu Kim , Myeung Ju Shin , Nari Kim , Song-Hee Lee , Young-Chul Kwon , Han Kyu Ko , Hyeon-Su Ro , Hyun-Sook Lee; J. Microbiol. 2013;51(1):118-122. Published online March 2, 2013; DOI: https://doi.org/10.1007/s12275-013-2351-2

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Abstract: A mycovirus was isolated from an edible mushroom, Lentinula edodes, that was suffering from a severe epidemic. Fractionation of the diseased cell extract by isopycnic centrifugation with 50% CsCl revealed that the diseased mushroom was infected by Lentinula edodes spherical virus (LeSV), a new spherical virus with a diameter of 55 nm. The particle of LeSV encapsidated the 12 kb RNA genome by a 120 kDa coat protein. BLAST analysis of the partially sequenced LeSV genome showed 95% sequence identity with a putative RNAdependent RNA polymerase (RdRp) gene of the mycovirus HKB, which was previously reported as being a doublestranded RNA (dsRNA) element. In contrast to HKB, the RNA genome in LeSV is encapsidated by the 120 kDa coat protein. To confirm that the LeSV coat protein is encoded by the viral genome, the N-terminal amino acid sequence of the coat protein was determined. The resulting N-terminal amino acid sequence, N-SALDVAPVVPELYFXXLEV-C, was found to be located in the middle of the HKB ORF1, suggesting that the LeSV coat protein was indeed encoded by the virus. To detect LeSV in L. edodes, a primer set targeting the RdRp gene was designed based on the partial sequence of the LeSV genome. RT-PCR analysis showed that 56 of the 84 commercially available dikaryotic cultivars carry LeSV. The transmission pattern of the virus was determined by analysing basidiospores from LeSV-infected and LeSVfree fruiting bodies. Nine out of 10 basidiospores from the LeSV-infected cultivars contained the virus while the spores from the LeSV-free parent were free of LeSV, suggesting that vertical transmission is the primary mode of LeSV propagation.

NOTE] The Microbial Population in the Air of Cultivation Facility of Oyster Mushrooms: Se Chul Chun , Yu Na Ahn , Sajid Mohamad Khan , Il Min Chung , Hyang Yoen Won , Chang Sung Jhune , Yool Jin Park; J. Microbiol. 2012;50(6):1053-1057. Published online December 30, 2012; DOI: https://doi.org/10.1007/s12275-012-2195-1

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Abstract: The microbial population in the air of mushroom cultivation facility was studied to understand the population structure and size depending on the cultivation methods and regions. The air contents of ten farmers’ oyster mushroom cultivation facilities in Kyunggi province were sampled. The results indicated that there was no difference in population size depending on the regions of mushroom cultivation. In addition, the population size of bacteria in the growth room was bigger than that of the cooling room and outside of the mushroom house, but the fungal population was similar in size between cultivation stages. With regard to population structure, Pseudomonas and Penicillium species were most frequently isolated from the air of oyster mushroom cultivation facility.

Screening, Purification, and Characterization of an Extracellular Prolyl Oligopeptidase from Coprinopsis clastophylla: Jen-Tao Chen , Mei-Li Chao , Chiou-Yen Wen , Wen-Shen Chu; J. Microbiol. 2012;50(4):652-659. Published online August 25, 2012; DOI: https://doi.org/10.1007/s12275-012-2099-0

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Abstract: Culture filtrates of 22 mushrooms were screened for extracellular prolyl oligopeptidase activity. Four strains with relatively high enzyme activity were all from inky cap mushrooms. The production of Coprinopsis clastophylla prolyl oligopeptidase was associated with the growth of the fungus and the enzyme was not released by cell lysis. The enzyme was purified 285-fold to a specific activity of 52.05 U/mg. It was purified to a single band on a native polyacrylamide gel. However, the enzyme separated into three bands on a sodium dodecyl sulfate-polyacrylamide gel with mobility corresponding to molecular weights of approximately 84, 60, and 26 kDa. The results of tandem mass spectrometric analysis revealed that the 60 kDa protein was likely a degradation product of the 84 kDa protein. The isoelectric point of the purified enzyme was 5.2. The purified enzyme had an optimal pH and temperature of 8.0 and 37°C, respectively. Diisopropyl fluorophosphate (DFP), p-chloromercuribenzoaic acid (PCMB), Hg2+, and Cu2+ strongly inhibited C. clastophylla prolyl oligopeptidase. This enzyme is a serine peptidase and one or more cysteine residues of the enzyme are close to the active site.

Journal Articles

Purification and Characterization of a Novel Laccase from the Edible Mushroom Hericium coralloides: Ya-Jie Zou , He-Xiang Wang , Tzi-Bun Ng , Chen-Yang Huang , Jin-Xia Zhang; J. Microbiol. 2012;50(1):72-78. Published online February 27, 2012; DOI: https://doi.org/10.1007/s12275-012-1372-6

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Abstract: A novel laccase from the edible mushroom Hericium coralloides was purified by ion exchange chromatography on diethylaminoethyl (DEAE) cellulose, carboxymethyl (CM) cellulose, and Q-Sepharose columns followed by fast protein liquid chromatography gel filtration on a Superdex 75 column. Analysis by gel filtration and SDS-PAGE indicated that the protein is a monomer in solution with a molecular mass of 65 kDa. Its N-terminal amino acid sequence was AVGDDTPQLY, which exhibits partial sequence homology to previously isolated laccases. Optimum activity was observed at pH 2.2 and at 40°C. The enzyme showed activity toward a variety of substrates, the most sensitive of which was 2,2􍿁-azinobis [3-ethylbenzothiazolone-6-sulfonic acid] diammonium salt (ABTS). The degradation activity toward substrates was ABTS > N,N-dimethyl-1,4-phenylenediamine > catechol > 2-methylcatechol > pyrogallol. The laccase did not exert any antiproliferative activity against Hep G2 or MCF 7 tumor cell lines at a concentration of 60 μM, unlike some previously reported mushroom proteins, but showed significant activity toward human immunodeficiency virus-1 (HIV-1) reverse transcriptase with an IC50 of 0.06 μM.

A Novel Ribonuclease with Potent HIV-1 Reverse Transcriptase Inhibitory Activity from Cultured Mushroom Schizophyllum commune: Yong-Chang Zhao , Guo-Qing Zhang , Tzi-Bun Ng , He-Xiang Wang; J. Microbiol. 2011;49(5):803-808. Published online November 9, 2011; DOI: https://doi.org/10.1007/s12275-011-1098-x

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Abstract: A 20-kDa ribonuclease (RNase) was purified from fresh fruiting bodies of cultured Schizophyllum commune mushrooms. The RNase was not adsorbed on Affi-gel blue gel but adsorbed on DEAE-cellulose and CM-cellulose. It exhibited maximal RNase activity at pH 6.0 and 70°C. It demonstrated the highest ribonucleolytic activity toward poly (U) (379.5 μ/mg), the second highest activity toward poly (C) (244.7 μ/mg), less activity toward poly (A) (167.4 μ/mg), and much weaker activity toward poly (G) (114.5 μ/mg). The RNase inhibited HIV-1 reverse transcriptase with an IC50 of 65 μM. No effect on [3H-methyl]-thymidine uptake by lymphoma MBL2 cells and leukemia L1210 cells was observed at 100 μM concentration of the RNase. A comparison of RNases from S. commune and Volvariella volvacea revealed that they demonstrated some similarities in N-terminal amino acid sequence, optimum pH and polyhomoribonucleotide specificity. However, some differences in chromatographic behavior and molecular mass were observed.

Research Support, Non-U.S. Gov'ts

NOTE] Initial Acidic pH Is Critical for Mycelial Cultures and Functional Exopolysaccharide Production of an Edible Mushroom, Laetiporus sulphureus var. miniatus JM 27: Min Jeong Seo , Min Jeong Kim , Hye Hyeon Lee , Sung Ryeal Kim , Byoung Won Kang , Jeong Uck Park , En Ju Rhu , Yung Hyun Choi , Yong Kee Jeong; J. Microbiol. 2010;48(6):881-884. Published online January 9, 2011; DOI: https://doi.org/10.1007/s12275-010-0410-5

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Abstract: We conducted a time course experiment on mycelial cultures of Laetiporus sulphureus var. miniatus. The strain showed significant survival in an initial pH range of 2.0 to 7.0 for 24 days, during which time oxalic acid was accumulated. A structural analysis of purified exopolysaccharide suggested that it contained 96.1% glucose, and the mode of linkage was mainly-4-Glcp-(1-units, with branches at the C-6 position consisting of a Glcp-(1-4) linked side chain. An exopolysaccharide purified from the acidophilic strain was added to cultured U937 cells, resulting in significantly increased transcription levels of p53 and p21 genes.

Purification and Biochemical Characterization of a 17 kDa Fibrinolytic Enzyme from Schizophyllum commune: In Suk Park , Jeong Uck Park , Min Jeong Seo , Min Jeong Kim , Hye Hyeon Lee , Sung Ryeal Kim , Byoung Won Kang , Yung Hyun Choi , Woo Hong Joo , Yong Kee Jeong; J. Microbiol. 2010;48(6):836-841. Published online January 9, 2011; DOI: https://doi.org/10.1007/s12275-010-0384-3

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Abstract: A fibrinolytic enzyme of the mushroom, Schizophyllum commune was purified with chromatographic methods, including a DEAE-Sephadex A-50 ion-exchange column and gel filtrations with Sephadex G-75 and Sephadex G-50 columns. The analysis of fibrin-zymography and SDS-PAGE showed that the enzyme was a monomeric subunit that was estimated to be approximately 17 kDa in size. The fibrinolytic activity of the enzyme in plasminogen-rich and plasminogen-free fibrin plates was 1.25 and 0.44 U/ml, respectively. The N-terminal amino acid sequence of the purified enzyme was identified as HYNIXNSWSSFID, which was highly distinguished from known fibrinolytic enzymes. The relative activity of the purified enzyme with an addition of 5 mM EDTA, Phosphoramidon, and Bestatin was about 76, 64, and 52%, respectively, indicating that it is a metalloprotease. The optimum temperature for the purified enzyme was approximately 45°C, and over 87% of the enzymatic activity was maintained as a stable state in a pH range from 4.0 to 6.0. Therefore, our results suggest that the potential thrombolytic agent from S. commune is a unique type of fibrinolytic enzyme.

Cloning and Sequence Analysis of a Glyceraldehyde-3-phosphate Dehydrogenase Gene from Ganoderma lucidum: Xu Fei , Ming Wen Zhao , Yu Xiang Li; J. Microbiol. 2006;44(5):515-522.; DOI: https://doi.org/2446 [pii]

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Abstract: A cDNA library of Ganoderma lucidum has been constructed using a Zap Express cloning vector. A glyceraldehyde-3-phosphate dehydrogenase gene (gpd) was isolated from this library by hybridization of the recombinant phage clones with a gpd-specific gene probe generated by PCR. By comparison of the cDNA and the genomic DNA sequences, it was found that the complete nucleotide sequence encodes a putative polypeptide chain of 338 amino acids interrupted by 6 introns. The predicted amino acid sequence of this gene shows a high degree of sequence similarity to the GPD proteins from yeast and filamentous fungi. The promoter region contains a CT-rich stretch, two CAAT boxes, and a consensus TATA box. The possibility of using the gpd promoter in the construction of new transformation vectors is discussed.

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