桑折 道済

Core-shell coagulates were prepared by hydrophobic heterocoagulation between non-charged poly(methyl methacrylate) (PMMA) core and cationically charged polystyrene (PSt) shell particles. Addition of NaCl and elevation of coagulation temperature to the vicinity of the glass transition temperature (T(g)) of the shell particles are found to be keys to successful heterocoagulation. Fluorescence resonance energy transfer (FRET) experiments using pyrene-labeled core and naphthalene-labeled shell particles showed that interdiffusion of the polymer chains at the interface between the core and shell particles is most effective when coagulates were prepared at or slightly above the T(g)s of both core and shell particles. Crown Copyright (C) 2010 Published by Elsevier B.V. All rights reserved.

Enzymatic glycosylation reactions by combined use of activated donor substrates and deactivated enzymes have been reviewed. Glycosidase-catalyzed transglycosylation reaction of sugar oxazolines to various glycosyl acceptors have successfully been achieved for the formation of poly- and oligo- saccharides such as chitin, glycosaminoglycans, and the core trisaccharide of N-linked oligosaccharide. These reactions proceeded efficiently using appropriate glycosidases under basic conditions, where the enzymes showed lower hydrolytic activity toward their natural substrates. In order to realize the transglycosylation reactions under neutral conditions, several mutants chitinaseA1 from Bacillus circulans WL-12 were constructed. Some of mutants were found to be efficient for transglycosylation under neutral conditions. These reactions could be applied to the stepwise synthesis of oligosaccharides and oligosaccharide-containing macromonomers. Novel thermosensitive polymers carrying oligosaccharides have successfully been synthesized by the radical copolymerization of the resulting macromonomers with N-isopropylacrylamide. One-pot chemo-enzymatic synthesis of oligosaccharides starting from a free sugar has been demonstrated via a sugar oxazoline intermediate formed in situ and the subsequent enzymatic transglycosylation. This novel technique of transferring an oligosaccharide moiety having an N-acetylhexosamine unit is a promising tool to pave the way to the new generation of oligosaccharide synthesis.

Several mutants of chitinase A1 from Bacillus circulans WL-12 have been prepared by site-directed mutagenesis of tryptophan 433. The hydrolytic activity of these mutants toward p-nitrophenyl chitobioside was lower than that of wild-type chitinase A1. The mutants were found to catalyze the transglycosylation reaction of the 1,2-oxazoline derivative of N-acetyllactosamine (GalGlcNAc-oxa) to N, N'-diacetylchitobiose (GlcNAc-GlcNAc), giving rise to a transglycosylated tetrasaccharide product (Gal-GlcNAc-GlcNAc-GlcNAc) in high yield. Based on these results, a one-pot synthesis of chitooli-gosaccharides by combined use of mutant W433Y and beta-galactosidase has been demonstrated. By controlling the reaction conditions, chitotriose and chitotetraose were obtained in 74% and 40% yield (determined by HPLC), respectively, without isolating any intermediate products.

A chitinolytic enzyme, chitinase A1 from Bacillus circulans WL-12, was found to catalyze a glycosyl-transferring reaction to form the N-linked oligosaccharide core structure, Man(beta 1-4)-GlcNAc(beta 1-4)-GlcNAc, by employing Man(beta 1-4)-GlcNAc-oxazoline as glycosyl donor. When the reaction was carried out in the presence of 20 v/v% acetone, the trisaccharide was obtained in 32% yield. It has been shown for the first time that a chitinase behaves like an endo-beta-N-acetylglucosaminidase in spite of low structural similarity between them.