Biotechnology for Biofuels and Bioproducts
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2024
·Vol. 17
·期 1
·1-17
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IF 3.3
Liu Lin,Wang Zhixing,Fang Yu,Yang Renfei,Pu Yi,Meng Xiangfeng,Liu Weifeng
Background The saprophytic filamentous fungus Trichoderma reesei represents one of the most prolific cellulase producers. The bulk production of lignocellulolytic enzymes by T. reesei not only relies on the efficient transcription of cellulase genes but also their efficient secretion after being translated. However, little attention has been paid to the functional roles of the involved secretory pathway in the high-level production of cellulases in T. reesei. Rab GTPases are key regulators in coordinating various vesicle trafficking associated with the eukaryotic secretory pathway. Specifically, Rab7 is a representative GTPase regulating the transition of the early endosome to the late endosome followed by its fusion to the vacuole as well as homotypic vacuole fusion. Although crosstalk between the endosomal/vacuolar pathway and the secretion pathway has been reported, the functional role of Rab7 in cellulase production in T. reesei remains unknown.Results A TrRab7 was identified and characterized in T. reesei. TrRab7 was shown to play important roles in T. reesei vegetative growth and vacuole morphology. Whereas knock-down of Trrab7 significantly compromised the induced production of T. reesei cellulases, overexpression of the key transcriptional activator, Xyr1, restored the production of cellulases in the Trrab7 knock-down strain (Ptcu-rab7KD) on glucose, indicating that the observed defective cellulase biosynthesis results from the compromised cellulase gene transcription. Down-regulation of Trrab7 was also found to make T. reesei more sensitive to various stresses including carbon starvation. Interestingly, overexpression of Snf1, a serine/threonine protein kinase known as an energetic sensor, partially restored the cellulase production of Ptcu-rab7KD on Avicel, implicating that TrRab7 is involved in an energetic adaptation to carbon starvation which contributes to the successful cellulase gene expression when T. reesei is transferred from glucose to cellulose.Conclusion sTrRab7 was shown to play important roles in T. reesei development and a stress response to carbon starvation resulting from nutrient shift. This adaptation may allow T. reesei to successfully initiate the inducing process leading to efficient cellulase production. The present study provides useful insights into the functional involvement of the endosomal/vacuolar pathway in T. reesei development and hydrolytic enzyme production.
研究领域:
微生物学
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分子生物学
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酶工程
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DOI
高影响因子
APPLIED BIOCHEMISTRY AND BIOTECHNOLOGY
·
2014
·Vol. 172
·期 7
·3448-3460
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IF 1.687
Zhang Jianfen,Chen Weiqing,Ke Wei,Chen Hong
The objective of this study was to screen glucoside 3-dehydrogenase (G3DH)-producing strain based on a high-throughput G3DH screening method. Optimization of culture conditions of the isolated strain was also applied in this study. This screening method employed electron transfer reaction in 96-well microtiter plates, α-methyl- d -glucoside, galactose, 2-deoxy- d -glucose, and 3- O -methyl- d -glucose were used as substrates. Using this screening method, one out of 78 strains isolated from different soil samples was obtained with high G3DH activity. The accuracy of the screening method was proved by alkaline treatment analysis of 3-keto sugars. The isolated strain was identified as Sphingobacterium faecium ZJF-D6 by phenotypic characterization and 16S rDNA sequence analysis. The culture conditions of S. faecium for G3DH production were optimized. Sucrose was found as the most suitable carbon source for the G3DH production. The highest G3DH production and cell growth were achieved using the medium at the initial pH of 7.0 at 25 °C for 36 h with activity of 8.03 × 10 −2 U/mL culture. This strain appears promising for potential application in the industry to produce 3-keto sugars. To our knowledge, this is the first report on S. faecium for G3DH production. The method described herein represents a useful tool for the high-throughput isolation of G3DH.
研究领域:
微生物学
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生物化学
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工业生物技术
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产品关联
DOI
PubMed
高影响因子
JOURNAL OF CHEMICAL TECHNOLOGY AND BIOTECHNOLOGY
·
2018
·Vol. 93
·期 11
·3135-3142
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IF 2.587
Lingqia Su,Tian Wang,Lingang Yu,Jing Wu
BACKGROUND We previously showed that extracellular production of naturally cytoplasmic proteins could be realized through co-expression with phospholipase C (PLC), which enhances cell membrane permeability through limited hydrolysis of membrane phospholipids. Leuconostoc mesenteroides sucrose phosphorylase (SP Lc ), a naturally cytoplasmic enzyme, exhibits excellent properties for synthesis of α-arbutin, but no systemic investigation of its preparation has thus far been reported. RESULTS Three PLCs were selected and expressed, without signal peptide, in Escherichia coli . More than 90% of the PLCs from Listeria monocytogenes and Bacillus cereus were present in the culture medium. When SP Lc was co-expressed with these PLCs, and the location of the PLC and SP Lc genes in the co-expression vector was optimized, the highest production was obtained when SP Lc gene was inserted upstream of the B. cereus PLC gene, as in BL21(DE3)/pETDuet -sp - bcplc . SP Lc production was then scaled up to a 3 L fermenter and fermentation conditions were optimized. When the initial medium contained 0.6 mmol L −1 ZnCl 2 and protein expression was induced at a dry cell weight of 15 g L −1 by the addition of lactose at 0.2 g L −1 h −1 at 30 °C, the extracellular SP Lc activity reached its highest level of 1445.1 U mL −1 This constituted 81.6% of total activity; the remainder of the enzyme (18.4%) remained inside the cells. CONCLUSION This is the first report describing extracellular expression of sucrose phosphorylase, and the highest yield ever reported was obtained. This provides the basis for industrial-scale production and application of sucrose phosphorylase. It also provides a potential method for improving the production of other proteins and fine chemicals. © 2018 Society of Chemical Industry
研究领域:
生物化学工程
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微生物学
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酶学
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生物技术
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产品关联
DOI
高影响因子
INTERNATIONAL JOURNAL OF MOLECULAR SCIENCES
·
2024
·Vol. 25
·期 19
·10547
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IF 4.9
Xiaona Zeng,Jiabei Song,Shengqiu Tang,Xiaoying Dong,Sheng Chen,Jie Kong,Liyi Chen,Yajuan Li,Guanming Shao,Yung-Hou Wong,Qingmei Xie
Tannins, one of the most common anti-nutritional factors in feed, can be effectively degraded by various enzymes secreted byAspergillus tubingensis(A. tubingensis). The cultivation method of fungi significantly impacts gene expression, which influences the production of enzymes and metabolites. In this study, we analyzed the tannin biodegredation efficiency and the transcriptomic responses ofA. tubingensisin liquid and solid cultures with tannin added. The observed morphology ofA. tubingensisresembled typical fungal hyphae of mycelium submerged and grown in liquid cultures, while mainly spore clusters were observed in solid cultures. Furthermore, the tannin biodegredation efficiency and protein secretion ofA. tubingensisin liquid cultures were significantly higher than in solid cultures. Additionally, 54.6% of the 11,248 differentially expressed genes were upregulated in liquid cultures, including AtWU_03490 (encoding ABC multidrug transporter), AtWU_03807 (ribonuclease III), AtWU_10270 (peptidyl-tRNA hydrolase), and AtWU_00075 (arabinogalactan endo-1,4-beta-galactosidase). Functional and gene ontology enrichment analyses indicated upregulation in processes including oxidation reduction, drug metabolism, and monocarboxylic acid metabolism. Overall, this study provides insight into the transcriptomic response to tannin biodegradation byA. tubingensisin different cultures and reveals that liquid cultures induce greater transcriptomic variability compared to solid cultures.
研究领域:
微生物学
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分子生物学
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生物技术
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基因组学
✓ 已验证
产品关联
DOI
PubMed
高影响因子
危险品化学品经营许可证（带存储）