Practical Preparation ofD-Galactosyl-β1→4-L-rhamnose Employing the Combined Action of Phosphorylases

Abstract

D-Galactosyl-1!4-L-rhamnose (GalRha) was produced enzymatically from 1.1 M sucrose and 1.0 M L-rhamnose by the concomitant actions of four enzymes (sucrose phosphorylase, UDP-glucose-hexose 1-phosphate uridylyltransferase, UDP-glucose 4-epimerase, and D-galactosyl-1!4-L-rhamnose phosphorylase) in the presence of 1.0 mM UDP-glucose and 30 mM inorganic phosphate. The accumulation of GalRha in 1 liter of the reaction mixture reached 230 g (the reaction yield was 71% from L-rhamnose). Sucrose and fructose in the reaction mixture were removed by yeast treatment, but isolation of GalRha by crystallization after yeast treatment was unsuccessful. Finally, 49 g of GalRha was isolated from part of the reaction mixture with yeast treatment by gel-filtration chromatography.

Diversity of phosphorylases in glycoside hydrolase families

“…The equilibrium is very valuable as it allows the use of sucrose as the starting material. In fact, several glycosides were prepared in excellent yield by the concerted actions of sucrose phosphorylase and another phosphorylase starting from sucrose (Kitaoka and Hayashi 2002;Kitaoka 2007, 2009;Nakajima et al 2010b;Nihira et al 2014a;Abe et al 2015).…”

Unique Sugar Metabolic Pathways of Bifidobacteria

“…106) D-Galactosyl-1,4-L-rhamnose was also produced using the corresponding phosphorylase. 107) Studies of the prebiotic effects of LNB on bifidobacteria in vitro 36,37) as well as its thermal decomposition mechanism 108) became possible only when the disaccharide became abundantly available due to this technology. LNB and GNB structures are found in biologically functional glycoconjugates, such as glycoproteins (O-glycans) and glycolipids (glycosphingolipids and blood type antigens).…”

2-O-α-D-Glucosylglycerol Phosphorylase from Bacillus selenitireducens MLS10 Possessing Hydrolytic Activity on β-D-Glucose 1-Phosphate

“…In addition, the reversibility allows the practical synthesis of oligosaccharides from the readily available natural sugars by using a single phosphorylase [7], [8], by the combined reaction of two phosphorylases that share the same sugar 1-phosphate or utilizing the compatible ones [1], [9]–[13], or by the combined reaction of two phosphorylases producing different sugar 1-phosphates with additional enzymes to convert the sugar 1-phosphates [14]–[16].…”