Abstract
Stabilization ponds are a common treatment technology for wastewater generated by dairy industries. Large proportions of cheese whey are thrown into these ponds, creating an environmental problem because of the large volume produced and the high biological and chemical oxygen demands. Due to its composition, mainly lactose and proteins, it can be considered as a raw material for value-added products, through physicochemical or enzymatic treatments. β-Galactosidases (EC 3.2.1.23) are lactose modifying enzymes that can transform lactose in free monomers, glucose and galactose, or galactooligosacharides. Here, the identification of novel genes encoding β-galactosidases, identified via whole-genome shotgun sequencing of the metagenome of dairy industries stabilization ponds is reported. The genes were selected based on the conservation of catalytic domains, comparing against the CAZy database, and focusing on families with β-galactosidases activity (GH1, GH2 and GH42). A total of 394 candidate genes were found, all belonging to bacterial species. From these candidates, 12 were selected to be cloned and expressed. A total of six enzymes were expressed, and five cleaved efficiently ortho-nitrophenyl-β-galactoside and lactose. The activity levels of one of these novel β-galactosidase was higher than other enzymes reported from functional metagenomics screening and higher than the only enzyme reported from sequence-based metagenomics. A group of novel mesophilic β-galactosidases from diary stabilization ponds’ metagenomes was successfully identified, cloned and expressed. These novel enzymes provide alternatives for the production of value-added products from dairy industries’ by-products.
Highlights
Whey is the main by-product of dairy industries
Several studies have been published in the past decades on the use of β-galactosidases from different sources for lactose hydrolysis [4,5,6]
With the improvement of sequencing technologies, screening uncultured microorganisms by sequence-based comparison to known sequences has been a useful approach for the identification of novel enzymes with potential for industrial applications [24,25,26]
Summary
Whey is the main by-product of dairy industries. It is composed mainly of lactose (4.5–5% w/v), proteins (0.6–0.8% w/v), lipids (0.4–0.5–5% w/v) and mineral salts (8–10% of dried extract). The estimated production of whey worldwide is about 180–190 million tons/year [1]. The high chemical and biological oxygen demands and the large volume of production make whey an important environmental problem. Β-Galactosidase (β-D-galactoside galactohydrolase or lactase; EC 3.2.1.23) is a type of glycoside hydrolase that has important industrial applications, such as lactose hydrolysis and transgalactosylation [2,3]. Several studies have been published in the past decades on the use of β-galactosidases from different sources for lactose hydrolysis [4,5,6]. Commercial enzymes were obtained from a limited number of species, including Aspergillus, Bacillus and Kluyveromyces
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