Duyệt theo Tác giả "Vu, Van Van"

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  • Hình ảnh thu nhỏ
    Tài liệu
    Cloning of AA9 Polysaccharide Monooxygenase gene AN3860 into pEX2B for expression in Aspergillus oryzae
    (Trường Đại học Nguyễn Tất Thành, 2020-06-29) Ngo, Thi Cam Nhung; Vu, Van Van
    Polysaccharide monooxygenases (PMOs) catalyze oxidative degradation of recalcitrant carbohydrate chains in cellulose, starch, and chitin. In biofuel industry, the conversion of rich lignocellulose source to fermentable sugars by hydrolytic cellulases can be synergistically boosted by cellulose-active PMOs (AA9 PMOs) found in a vast number of fungi that grow on biomass. Aspergillus nidulans, a filamentous fungus, possess a dozen of PMO-encoding genes, but only the AN3860 is expressed at a high level when cultured with wheat straw as the sole carbon source. Bioinformatic analysis indicates that AN3860 belongs to type 3 AA9 PMO subfamily that is capable of hydroxylating both C1 and C4 of the glycosidic linkages. Therefore, AN3860 may be a potential enzyme to improve cellulose hydrolysis efficiency, which has not been characterized. In this study, we describe the AN3860 cloning into Aspergillus oryzae AUT1-PlD. To facilitate the purification of AN3860, we added a CBM20 tag to its C-terminal. The recombinant vector was designed and constructed successfully. Simultaneously, we have obtained the clone of A. oryzae carrying the target gene by the ATMT method. Further expression optimization and characterization of AN3860 by both activity assays and spectroscopic techniques are underway.
  • Hình ảnh thu nhỏ
    Tài liệu
    Cloning of an AA13 Polysaccharide Monooxygenase Gene from Neurospora crassa into the plasmid pEX2B
    (Trường Đại học Nguyễn Tất Thành, 2020) Ho, Ta Giap; Ngo, Nguyen Vu; Vu, Van Van
    Polysaccharide monooxygenases (PMOs) cleave glycoside linkages in polysaccharides, which has great potential in manufacturing biofuel from polysaccharide biomasses. Therefore, heterologous production of PMOs is required for the development of their application. We aim to clone PMO genes into the plasmid pEX2B to express PMOs in Aspergillus oryzae, a filamentous fungus widely used for industrial enzyme production. Here we cloned the NCU08746 gene, an AA13 PMO from Neurospora crassa, into pEX2B. We successfully designed primers and amplified the NCU08746 gene from the N. crassa genome. NCU08746 was then cloned into the pEX2B vector. The recombinant plasmid pEX2B-NCU08746 was subsequently transferred into A. tumefaciens AGL1 prior to being transferred into A. oryzae. This work provides the foundation for further study on PMOs expression in A. oryzae.
  • Hình ảnh thu nhỏ
    Tài liệu
    Recombinant protein expression of Magnaporthe oryzae MGG06069 and its polysaccharide monooxygenase domain in Escherichia coli
    (Nguyen Tat Thanh University, 2022) Bui, Cong Chinh; Ho, Ta Giap; Vu, Van Van
    MGG06069 is a polysaccharide monooxygenase protein that originated from the fungus that causes rice blast, Magnaporthe oryzae. It has the potential to speed up the cellulolysis process, which can be beneficial for industry. In this study, the full-length MGG06069 protein and its polysaccharide monooxygenase domain were expressed in Escherichia coli in order to produce recombinant proteins on a large scale for further enzymatic experiment. It was shown that the amount of the recombinant polysaccharide monooxygenase domain protein was much higher compared to that of the full-length protein. In addition, only one band of expected size was obtained for the recombinant PMO domain protein. These results suggest the purified polysaccharide monooxygenase domain of MGG06069 that can be used for enzymatic assays in the future.
  • Hình ảnh thu nhỏ
    Tài liệu
    Study on AA10 expression in E. coli
    (Trường Đại học Nguyễn Tất Thành, 2021) Vu, Van Van; Ngo, Thi Cam Nhung
    The GlcNAc-binding protein A (GbpA) has been known as a virulent factor of Vibrio vulnificus pathogen. Domain 1 of GbpA adhesion takes responsibility of binding both human intestine and the chitinous surface. The domain 1 structure is similar to a polysaccharide monooxygenase (PMO) AA10-type (PMO), which catalyzed oxidation toward the recalcitrant chitin polymer. The role of the VvPMO10 module in catalytic functions has not been fulfilled characterized. To aim of the VvPMO10 study, this protein was cloned to the pET22b system and transformed into the E. coli BL21 (DE3) strain. The recombinant enzyme was expressed at 37 0C with IPTG induced. Total protein was checked by SDS-PAGE method and stained using Coomassie blue solution. The target band showed a band of 20 kDa as expectation. Thus, the heterologous protein was expressed successfully in E. coli BL21 (DE3) strain and becomes the materials for future study.