49 天然多糖素材からのエリシター活性オリゴ糖の作出(口頭発表の部)

抄録

Elicitor-active oligosaccharides have been produced from natural polysaccharides by the two approaches, 1) partial hydrolysis of the polysaccharides, 2) enzymatic synthesis utilizing glycosidase-catalyzed transglycosylation reaction. Natural polysaccharides were examined for their activity as elicitors in alfalfa cotyledons. Laminaran, which had a marked elicitor activity, was partially hydrolyzed and then purified by charcoal and gel filtration columns. Introduction of the pyridylamino group into the elicitor-active oligosaccharides was attempted in order to facilitate isolation. Their HPLC analysis showed three main peaks. The individual peaks (LN-1, 2, 3) were collected and subjected to the alfalfa cotyledon assay. LN-3 showed the highest activity (MEC; 650 nM). Based on the analyses of glycosyl-linkage and fragments by acetolysis, LN-3 is a linear pyridylaminated hepta-β-glucoside which consists of a PA-laminaribiose unit at the reducing end, a glucose and two laminaribiose units with three β-(1→6)-linkages. The glycosyl-sequence analysis only allows us to depict the structure as shown in Fig. 1-4.. Chitooligosaccharides and partially N-acetylchitooligosaccharides were synthesized by lysozyme-catalyzed transglycosylation reaction using N-acetylchitotriose and N-monochloroacetylchitotriose as substrates. The signals due to the H-1 protons of the GlcN units at δ4.85-4.95 and those of the GlcNAc units at δ4.55-4.65 in the ^1H NMR spectra indicated that glycosidic bond formation occurred in a regio- and stereoselective manner to afford oligosaccharides having β-(1→4) -linkage. The MALDI TOF MS analysis revealed that the range of the maximum molecular weight is 1700-2600 (d.p. 10-14). The chitooligosaccharide and partially N-acetylchitooligosaccharide (d.a. 36%) synthesized in an ammonium sulfate-containing buffer exhibited strong activity for phytoalexin (+)-pisatin induction in pea epicotyl elicitor assay. The transglycosylation of N-monochloroacetylated sugars followed by N-dechloroacetylation could be one of the most effective methods for producing chitooligosaccharides and partially N-acetylchitooligosaccharides, which could possess a variety of biological activities.