Homologous expression of Aspergillus niger α-L-rhamnosidase and its application in enzymatic debittering of Ougan juice.

IF 2 4区生物学 Q3 BIOTECHNOLOGY & APPLIED MICROBIOLOGY

Biotechnology Letters Pub Date : 2024-09-05 DOI:10.1007/s10529-024-03531-x

Fei Zhang, Xue Wang, Lixia Pan, Zhao Wang, Jianyong Zheng

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Abstract

The α-L-rhamnosidase (rha1) gene was homologously expressed in Aspergillus niger strains CCTCC 206047 and CCTCC 206047ΔpyrG, using hygromycin B and auxotrophic as selection markers. The engineered A. niger strains RHA001-1 and RHA003-1 were screened, yielding α-L-rhamnosidase activities of 20.81 ± 0.56 U/mL and 15.35 ± 0.87 U/mL, respectively. The copy numbers of the rha1 gene in strains RHA001-1 and RHA003-1 were found to be 18 and 14, respectively. Correlation analysis between copy number and enzyme activity in the A. niger strains revealed that α-L-rhamnosidase activity increased with the copy number of the rha1 gene. Recombinant α-L-rhamnosidase was utilized for the enzymatic debittering of Ougan juice, and its process conditions were optimized. Furthermore, the primary bitter substance neohesperidin (2.22 g/L) in Ougan juice was converted into hesperetin 7-O-glucoside (1.47 g/L) and hesperidin (0.143 g/L). This study presents a novel approach for the production of food-grade α-L-rhamnosidase and establishes a technical foundation for its application in the beverage industry.

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黑曲霉α-L-鼠李糖酶的同源表达及其在乌干果汁酶解脱盐中的应用。

以潮霉素 B 和辅助营养型为选择标记，在黑曲霉菌株 CCTCC 206047 和 CCTCC 206047ΔpyrG 中同源表达了 α-L 鼠李糖酶（rha1）基因。筛选出的黑曲霉工程菌株RHA001-1和RHA003-1的α-L-鼠李糖酶活性分别为20.81 ± 0.56 U/mL和15.35 ± 0.87 U/mL。RHA001-1 和 RHA003-1 菌株中 rha1 基因的拷贝数分别为 18 和 14。黑曲霉菌株中拷贝数与酶活性之间的相关性分析表明，α-L-鼠李糖酶活性随 rha1 基因拷贝数的增加而增加。重组α-L-鼠李糖酶被用于乌干果汁的酶法脱苦，其工艺条件也得到了优化。此外，乌干果汁中的主要苦味物质新橙皮甙（2.22 克/升）被转化为橙皮甙 7-O-葡萄糖苷（1.47 克/升）和橙皮甙（0.143 克/升）。这项研究提出了一种生产食品级α-L-鼠李糖酶的新方法，并为其在饮料行业的应用奠定了技术基础。

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来源期刊

Biotechnology Letters 工程技术-生物工程与应用微生物

CiteScore

5.90

自引率

3.70%

发文量

108

审稿时长

1.2 months

期刊介绍： Biotechnology Letters is the world’s leading rapid-publication primary journal dedicated to biotechnology as a whole – that is to topics relating to actual or potential applications of biological reactions affected by microbial, plant or animal cells and biocatalysts derived from them. All relevant aspects of molecular biology, genetics and cell biochemistry, of process and reactor design, of pre- and post-treatment steps, and of manufacturing or service operations are therefore included. Contributions from industrial and academic laboratories are equally welcome. We also welcome contributions covering biotechnological aspects of regenerative medicine and biomaterials and also cancer biotechnology. Criteria for the acceptance of papers relate to our aim of publishing useful and informative results that will be of value to other workers in related fields. The emphasis is very much on novelty and immediacy in order to justify rapid publication of authors’ results. It should be noted, however, that we do not normally publish papers (but this is not absolute) that deal with unidentified consortia of microorganisms (e.g. as in activated sludge) as these results may not be easily reproducible in other laboratories. Papers describing the isolation and identification of microorganisms are not regarded as appropriate but such information can be appended as supporting information to a paper. Papers dealing with simple process development are usually considered to lack sufficient novelty or interest to warrant publication.