Novel Enzyme for Synthesizing β-1,2-Galactooligosaccharides

Carbohydrate chains, or glycans, are complex sugar-based molecules essential for various biological functions and structures in the human body. Among them, galactosides form a distinct category found in plants, animals, and microorganisms.

Galactosides play key roles in nature. In plants, they contribute to cell wall integrity, while in human nutrition, they are present in prebiotic oligosaccharides that promote gut health. Additionally, glycans containing galactose are often used in processed foods such as juice and powdered milk due to their potential health benefits.

Understanding the enzymes that break down these glycans is crucial for exploring their prebiotic properties and enhancing their applications in food and health products. One such enzyme group, β-galactosidases, releases galactose from galactosides. Different β-galactosidases target specific galactosides and are found in mammals, including human gut bacteria like Bifidobacterium, which aids in digesting complex carbohydrates.

Recent research has highlighted the capabilities of another gut bacterium, Bacteroides xylanisolvens, known for its ability to metabolize a diverse range of carbohydrates. However, its precise functions remain largely unexplored.

In a groundbreaking study, a research team led by Associate Professor Masahiro Nakajima from the Tokyo University of Science (TUS) identified a novel β-galactosidase enzyme in B. xylanisolvens. This enzyme specifically targets unique galactose-containing glycans that may have prebiotic properties. The study, published in Communications Biology, included contributions from Mr. Yutaka Nakazawa (TUS), Associate Professor Hiroyuki Nakai (Niigata University), and Assistant Professor Tomohiko Matsuzawa (Kagawa University).

“Many glycans have complex structures with unknown functions and potential uses,” said Dr. Nakajima. “Since enzymes are essential for glycan synthesis, discovering new ones is extremely important. Our novel enzyme could enable large-scale production of unique glycans with prebiotic potential, benefiting human health.”

B. xylanisolvens contains multiple genes encoding β-galactosidases. The researchers identified one gene, Bxy_22780, responsible for encoding a previously unknown β-galactosidase. Initially, this enzyme showed no activity on natural β-galactosides. However, in experiments using a nucleophile mutant, α-D-galactosyl fluoride (α-GalF) as a donor substrate, and galactose or D-fucose as an acceptor substrate, the team successfully detected reaction products. Nuclear magnetic resonance studies confirmed the formation of β-1,2-galactobiose, a disaccharide.

Further analysis revealed that the Bxy_22780 enzyme is highly selective for galactooligosaccharides (GOS), particularly those with β-1,2-galactosidic linkages. Kinetic studies confirmed its effectiveness in breaking down β-1,2-galactobiose and β-1,2-galactotriose. To determine the basis of its specificity, the researchers conducted X-ray diffraction studies, which showed that the enzyme binds methyl β-galactopyranose at a critical site known as subsite +1, optimally positioning the molecule for breaking down these specific sugar chains.

Nakajima emphasized the significance of these findings: “β-1,2-Galactooligosaccharides and their associated enzymes have been rarely studied. Our discovery is a crucial step in understanding these unique glycans, whose functions remain largely unknown.”

While the prebiotic properties of β-1,2-galactooligosaccharides have yet to be confirmed, they hold strong potential. Moreover, this enzyme could pave the way for new therapeutic strategies. For instance, Chagas disease—a life-threatening illness caused by a parasite producing glycans with β-1,2-galactosidic linkages—could potentially be targeted through enzyme-based treatments.

The discovery of Bxy_22780 marks a significant advancement in prebiotic research, opening doors to innovative food and health applications. This enzyme could aid in the development of next-generation prebiotic products designed to enhance gut health, improve digestion, and even contribute to new medical treatments. As research continues, its potential to transform the food and pharmaceutical industries becomes increasingly promising.