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Immobilization of β-Galactosidases from Lactobacillus on Chitin Using a Chitin-Binding Domain

Author:
Pham, Mai-Lan, Leister, Tatjana, Nguyen, Hoang Anh, Do, Bien-Cuong, Pham, Anh-Tuan, Haltrich, Dietmar, Yamabhai, Montarop, Nguyen, Thu-Ha, Nguyen, Tien-Thanh
Source:
Journal of agricultural and food chemistry 2017 v.65 no.14 pp. 2965-2976
ISSN:
0021-8561
Subject:
Lactobacillus delbrueckii subsp. bulgaricus, Lactobacillus plantarum, Lactobacillus reuteri, active sites, beta-galactosidase, carbohydrate binding, chitin, immobilized enzymes, lactose
Abstract:
Two β-galactosidases from Lactobacillus, including a heterodimeric LacLM type enzyme from Lactobacillus reuteri L103 and a homodimeric LacZ type β-galactosidase from Lactobacillus bulgaricus DSM 20081, were studied for immobilization on chitin using a carbohydrate-binding domain (chitin-binding domain, ChBD) from a chitinolytic enzyme. Three recombinant enzymes, namely, LacLM-ChBD, ChBD-LacLM, and LacZ-ChBD, were constructed and successfully expressed in Lactobacillus plantarum WCFS1. Depending on the structure of the enzymes, either homodimeric or heterodimeric, as well as the positioning of the chitin-binding domain in relation to the catalytic domains, that is, upstream or downstream of the main protein, the expression in the host strain and the immobilization on chitin beads were different. Most constructs showed a high specificity for the chitin in immobilization studies; thus, a one-step immobilizing procedure could be performed to achieve up to 100% yield of immobilization without the requirement of prior purification of the enzyme. The immobilized-on-chitin enzymes were shown to be more stable than the corresponding native enzymes; especially the immobilized LacZ from L. bulgaricus DSM20081 could retain 50% of its activity when incubated at 37 °C for 48 days. Furthermore, the immobilized enzymes could be recycled for conversion up to eight times with the converting ability maintained at 80%. These results show the high potential for application of these immobilized enzymes in lactose conversion on an industrial scale.
Agid:
5799690