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Transglycosylating β-d-galactosidase and α-l-fucosidase from Paenibacillus sp. 3179 from a hot spring in East Greenland.
MicrobiologyOpen ( IF 3.9 ) Pub Date : 2019-12-23 , DOI: 10.1002/mbo3.980
Mariane S Thøgersen 1 , Stefan J Christensen 1 , Morten Jepsen 1 , Lars H Pedersen 2 , Peter Stougaard 1
Affiliation  

Thermal springs are excellent locations for discovery of thermostable microorganisms and enzymes. In this study, we identify a novel thermotolerant bacterial strain related to Paenibacillus dendritiformis, denoted Paenibacillus sp. 3179, which was isolated from a thermal spring in East Greenland. A functional expression library of the strain was constructed, and the library screened for β‐d‐galactosidase and α‐l‐fucosidase activities on chromogenic substrates. This identified two genes encoding a β‐d‐galactosidase and an α‐l‐fucosidase, respectively. The enzymes were recombinantly expressed, purified, and characterized using oNPG (2‐nitrophenyl‐β‐d‐galactopyranoside) and pNP‐fucose (4‐nitrophenyl‐α‐l‐fucopyranoside), respectively. The enzymes were shown to have optimal activity at 50°C and pH 7–8, and they were able to hydrolyze as well as transglycosylate natural carbohydrates. The transglycosylation activities were investigated using TLC and HPLC, and the β‐d‐galactosidase was shown to produce the galactooligosaccharides (GOS) 6'‐O‐galactosyllactose and 3'‐O‐galactosyllactose using lactose as substrate, whereas the α‐l‐fucosidase was able to transfer the fucose moiety from pNP‐fuc to lactose, thereby forming 2'‐O‐fucosyllactose. Since enzymes that are able to transglycosylate carbohydrates at elevated temperature are desirable in many industrial processes, including food and dairy production, we foresee the potential use of enzymes from Paenibacillus sp. 3179 in the production of, for example, instant formula.

中文翻译:


来自类芽孢杆菌属的转糖基化 β-d-半乳糖苷酶和 α-l-岩藻糖苷酶。 3179 来自东格陵兰岛的温泉。



温泉是发现热稳定性微生物和酶的绝佳场所。在这项研究中,我们鉴定了一种与树状类芽孢杆菌相关的新型耐热细菌菌株,称为类芽孢杆菌sp。 3179,从东格陵兰岛的一个温泉中分离出来。构建了该菌株的功能表达文库,并筛选了该文库在显色底物上的β- d-半乳糖苷酶和α -l-岩藻糖苷酶活性。这鉴定了两个分别编码 β- d-半乳糖苷酶和 α- l-岩藻糖苷酶的基因。分别使用o NPG(2-硝基苯基-β- d-吡喃半乳糖苷)和p NP-岩藻糖(4-硝基苯基-α- l-吡喃岩藻糖苷)对酶进行重组表达、纯化和表征。这些酶在 50°C 和 pH 7-8 时具有最佳活性,并且能够水解和转糖基化天然碳水化合物。使用 TLC 和 HPLC 研究转糖基活性,结果显示 β- d-半乳糖苷酶以乳糖为底物产生低聚半乳糖 (GOS) 6'- O-半乳糖基乳糖和 3'- O-半乳糖基乳糖,而 α- l-岩藻糖苷酶能够将岩藻糖部分从p NP-fuc 转移到乳糖,从而形成 2'- O-岩藻糖基乳糖。由于能够在高温下转糖基化碳水化合物的酶在许多工业过程(包括食品和乳制品生产)中是理想的,因此我们预见来自类芽孢杆菌属的酶的潜在用途。 3179 用于生产速溶配方奶粉等。
更新日期:2019-12-23
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