Screening of a genome-reduced Corynebacterium glutamicum strain library for improved heterologous cutinase secretion

IF 4.8 2区 生物学 Q1 BIOTECHNOLOGY & APPLIED MICROBIOLOGY
Johannes Hemmerich, Mohamed Labib, Carmen Steffens, Sebastian J. Reich, Marc Weiske, Meike Baumgart, Christian Rückert, Matthias Ruwe, Daniel Siebert, Volker F. Wendisch, J?rn Kalinowski, Wolfgang Wiechert, Marco Oldiges
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引用次数: 15

Abstract

The construction of microbial platform organisms by means of genome reduction is an ongoing topic in biotechnology. In this study, we investigated whether the deletion of single or multiple gene clusters has a positive effect on the secretion of cutinase from Fusarium solani pisi in the industrial workhorse Corynebacterium glutamicum. A total of 22 genome-reduced strain variants were compared applying two Sec signal peptides from Bacillus subtilis. High-throughput phenotyping using robotics-integrated microbioreactor technology with automated harvesting revealed distinct cutinase secretion performance for a specific combination of signal peptide and genomic deletions. The biomass-specific cutinase yield for strain GRS41_51_NprE was increased by ~ 200%, although the growth rate was reduced by ~ 60%. Importantly, the causative deletions of genomic clusters cg2801-cg2828 and rrnC-cg3298 could not have been inferred a priori. Strikingly, bioreactor fed-batch cultivations at controlled growth rates resulted in a complete reversal of the screening results, with the cutinase yield for strain GRS41_51_NprE dropping by ~ 25% compared to the reference strain. Thus, the choice of bioprocess conditions may turn a ‘high-performance’ strain from batch screening into a ‘low-performance’ strain in fed-batch cultivation. In conclusion, future studies are needed in order to understand metabolic adaptations of C. glutamicum to both genomic deletions and different bioprocess conditions.

Abstract Image

改良异源角质酶分泌的谷氨酸棒状杆菌基因组还原菌株文库的筛选
利用基因组还原技术构建微生物平台生物一直是生物技术研究的热点。在这项研究中,我们研究了单个或多个基因簇的缺失是否会对工业主力谷氨酸棒状杆菌中茄枯镰刀菌的表皮酶分泌产生积极影响。利用枯草芽孢杆菌的两种Sec信号肽对22个基因组减少的菌株进行了比较。利用机器人集成微生物反应器技术和自动采集技术进行高通量表型分析,揭示了信号肽和基因组缺失的特定组合的独特角质酶分泌性能。菌株GRS41_51_NprE的生物量特异性角质酶产量提高了~ 200%,但生长速率降低了~ 60%。重要的是,基因组簇cg2801-cg2828和rrnC-cg3298的致病缺失不能被先验推断。引人注目的是,在控制生长速率的生物反应器中进行间歇培养,结果与筛选结果完全相反,菌株GRS41_51_NprE的表皮酶产量比参考菌株下降了~ 25%。因此,生物工艺条件的选择可能会将分批筛选的“高性能”菌株转变为分批培养的“低性能”菌株。总之,需要进一步的研究来了解谷氨酰胺对基因组缺失和不同生物过程条件的代谢适应。
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来源期刊
Microbial Biotechnology
Microbial Biotechnology BIOTECHNOLOGY & APPLIED MICROBIOLOGY-MICROBIOLOGY
CiteScore
9.80
自引率
3.50%
发文量
162
审稿时长
6-12 weeks
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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