Improvement of β-Xylosidase and Endoxylanase Activities in Talaromyces amestolkiae by Genetic Manipulation of the Transcriptional Activator XlnR

IF 5.7 2区 生物学
Ana Pozo-Rodríguez, Miguel Ángel Peñalva, Jorge Barriuso, Eduardo A. Espeso, María Jesús Martínez
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引用次数: 0

Abstract

The ascomycete Talaromyces amestolkiae is a promising source of glycosyl hydrolases for hemicellulose degradation, as it contains a considerably higher number of genes encoding these enzymes than other fungi exploited for plant biomass valorisation. The development of genetic engineering tools could further improve its biotechnological potential. We report here a transformation system for T. amestolkiae based on pyrimidine auxotrophy complementation, which was used to successfully introduce both integrative and autonomously replicating plasmids. Then, we applied this tool to force the expression of the transcriptional activator XlnR, generating an engineered strain with enhanced β-xylosidase (1.4-fold) and endoxylanase (2.0-fold) activities compared to the wild-type cultured on xylan. Markedly larger improvements were obtained after introducing Ala788Val or Val785Phe substitutions in XlnR, achieving 3.3-fold and 3.9-fold increases in β-xylosidase and endoxylanase activities, respectively, in the case of XlnRV785F. This recombinant strain also displays a partial deregulation of the hemicellulolytic system when cultivated on glucose and glycerol (a low-cost and renewable substrate), yielding notably higher production of β-xylosidases (16.9-fold and 13.8-fold) and endoxylanases (31.9-fold and 22.7-fold) than the wild-type. Increased efficiencies of XlnRV785F enzymatic crudes in xylan saccharifications showed the potential of XlnR engineering to develop robust T. amestolkiae strains for the valorisation of hemicellulosic residues.

转录激活子XlnR的遗传修饰对阿梅塔芳酵母β-木糖苷酶和内木聚糖酶活性的影响
与其他用于植物生物量增殖的真菌相比,子囊菌Talaromyces amestolkiae含有大量编码这些酶的基因,因此它是用于半纤维素降解的糖基水解酶的有希望的来源。基因工程工具的发展可以进一步提高其生物技术潜力。在此,我们报道了一种基于嘧啶互补的amestolkiae转化系统,该系统成功地引入了整合质粒和自主复制质粒。然后,我们利用该工具强制表达转录激活子XlnR,产生了与在木聚糖上培养的野生型相比,β-木糖苷酶(1.4倍)和内木聚糖酶(2.0倍)活性增强的工程菌株。在XlnR中引入Ala788Val或Val785Phe替代后,XlnRV785F的β-木糖苷酶和内木聚糖酶活性分别提高了3.3倍和3.9倍,得到了更大的改善。当在葡萄糖和甘油(一种低成本和可再生的底物)上培养时,该重组菌株也显示出半纤维素分解系统的部分放松,其β-木糖苷酶(16.9倍和13.8倍)和内木聚糖酶(31.9倍和22.7倍)的产量明显高于野生型。XlnRV785F酶原在木聚糖糖化过程中的效率提高,表明XlnR工程有潜力开发出强大的amestolkiae菌株,用于半纤维素残基的增殖。
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来源期刊
Microbial Biotechnology
Microbial Biotechnology Immunology and Microbiology-Applied Microbiology and Biotechnology
CiteScore
11.20
自引率
3.50%
发文量
162
审稿时长
1 months
期刊介绍: Microbial Biotechnology publishes papers of original research reporting significant advances in any aspect of microbial applications, including, but not limited to biotechnologies related to: Green chemistry; Primary metabolites; Food, beverages and supplements; Secondary metabolites and natural products; Pharmaceuticals; Diagnostics; Agriculture; Bioenergy; Biomining, including oil recovery and processing; Bioremediation; Biopolymers, biomaterials; Bionanotechnology; Biosurfactants and bioemulsifiers; Compatible solutes and bioprotectants; Biosensors, monitoring systems, quantitative microbial risk assessment; Technology development; Protein engineering; Functional genomics; Metabolic engineering; Metabolic design; Systems analysis, modelling; Process engineering; Biologically-based analytical methods; Microbially-based strategies in public health; Microbially-based strategies to influence global processes
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