Enzymatic synthesis of 4-methylumbelliferyl (1→3)-β-d-glucooligosaccharides—new substrates for β-1,3-1,4-d-glucanase Original Research Article

The transglycosylation reactions catalyzed by β-1,3-d-glucanases (laminaranases) were used to synthesize a number of 4-methylumbelliferyl (MeUmb) (1→3)-β-d-gluco-oligosaccharides having the common structure [β-d-Glcp-(1→3)]n-β-d-Glcp-MeUmb, where n=1–5. The β-1,3-d-glucanases used were purified from the culture liquid of Oerskovia sp. and from a homogenate of the marine mollusc Spisula sachalinensis. Laminaran and curdlan were used as (1→3)-β-d-glucan donor substrates, while MeUmb-β-d-glucoside (MeUmbGlcp) was employed as a transglycosylation acceptor. Modification of [β-d-Glcp-(1→3)]2-β-d-Glcp-MeUmb (MeUmbG3) gives 4,6-O-benzylidene-d-glucopyranosyl or 4,6-O-ethylidene-d-glucopyranosyl groups at the non-reducing end of artificial oligosaccharides. The structures of all oligosaccharides obtained were solved by 1H and 13C NMR spectroscopy and electrospray tandem mass spectrometry. The synthetic oligosaccharides were shown to be substrates for a β-1,3-1,4-d-glucanase from Rhodothermus marinus, which releases MeUmb from β-di- and β-triglucosides and from acetal-protected β-triglucosides. When acting upon substrates with d.p.>3, the enzyme exhibits an endolytic activity, primarily cleaving off MeUmbGlcp and MeUmbG2.