Novel Synthesis of Lactosyl β-Glycosides and Analogs through Cellulase-mediated Transglycosylation or Condensation Reaction

Abstract

Alkyl β-lactosides were directly synthesized by β-lactosyl transfer reaction from p-nitrophenyl β-lactoside (Lacβ-pNP) to various 1-alkanols (n=2-12), utilizing a commercially available cellulase preparation of Trichoderma reesei C1. When 1-octanol and dodecanol were acceptors, octyl β-lactoside and dodecyl β-lactoside were obtained as transfer products, respectively. Furthermore, the enzyme catalyzed N-acetyllactosaminyl transfer reaction from p-nitrophenyl β-N-acetyllactosaminide (LacNAcβ-pNP) not only to 1-alkanol, but also to the OH-4 position of Man and Glc to produce the trisaccharides. As an alternative method, alkyl β-lactosides were synthesized by condensation reaction of lactose with alkanols. When various 1-alkanols (n: 2-8) were used, the corresponding alkyl β-lactosides were obtained in the yields of 1-4% based on lactose added. Further, condensation reaction between lactose and glycerol was effectively catalyzed to produce 1-O-β-lactosyl-(R,S)-glycerols and 2-O-β-lactosyl glycerol in a molar ratio of 7:3 and in a 20% yield based on lactose added. The enzyme also catalyzed the condensation with 2-propanol and allyl alcohol. In a similar manner, when LacNAc was used as a glycon substrate, the enzyme catalyzed the condensation with various alkanols and glycerol. The yields were lower than those obtained from reactions with lactose. Finally, purification of an enzyme possessing condensation activity was carried out by chromatographies using DEAE-Sepharose and Mono P columns, in order to reveal whether the same enzyme molecule would catalyze in both the lactosyl- and N-acetyllactosaminyl-condensing reactions. It was specified that a single enzyme works both condensation reactions, based on the substrate competition assay on hydrolysis.