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The beta-fructofuranosidase from the yeast Xanthophyllomyces dendrorhous (Xd-INV) catalyzes the synthesis of neo-fructooligosaccharides (neo-FOS of the 6G-series), which contain a beta(2-6) linkage between a fructose and the glucosyl moiety of sucrose. In this work we demonstrate that the enzyme is also able to fructosylate other carbohydrates that contain glucose, in particular disaccharides (maltose, isomaltulose, isomaltose, trehalose) and higher oligosaccharides (maltotriose, raffinose, maltotetraose), but not monosaccharides (glucose, fructose, galactose). With maltose as acceptor, the reaction in the presence of Xd-INV proceeded with high regioselectivity; the product was purified and chemically characterized, and turned out to be 6’-O--fructosylmaltose (neo-erlose). Using 100 g/L sucrose as fructosyl donor and 300 g/L maltose as acceptor, the maximum concentration of neo-erlose was 38.3 g/L. Thus, novel hetero-fructooligosaccharides with potential applications in the functional food and pharmaceutical industries can be obtained with Xd-INV., Projects BIO2010-20508-C04-01 and BIO2010-20508-C04-04 from Spanish Ministry of Science and Innovation supported this research. We thank Fundación Ramon Areces for an institutional grant to the Centro de Biología Molecular Severo Ochoa.

Different filamentous fungi isolated from molasses and jams (kiwi and fig) were screened for fructooligosaccharides (FOS) producing activity. Two strains, identified as Penicilium sizovae (CK1) and Cladosporium cladosporioides (CF215), were selected on the basis of the FOS yield and kestose/nystose ratio. In both strains the activity was mostly mycelium-bound. Starting from 600 g/L of sucrose, maximum FOS yield was 184 and 339 g/L for P. sizovae and C. cladosporioides, respectively. Interestingly, the highest FOS concentration with C. cladosporioides was reached at 93% sucrose conversion, which indicated a notable transglycosylation to hydrolysis ratio. The main FOS in the reaction mixtures were identified by HPAEC-PAD chromatography. C. cladosporioides synthesized mainly 1-kestose (158 g/L), nystose (97 g/L), 1F-fructosylnystose (19 g/L), 6-kestose (12 g/L), neokestose (10 g/L) and a disaccharide (34 g/L) that after its purification and NMR analysis was identified as blastose [Fru-beta(2-6)-Glc]. P. sizovae was very selective for the formation of 1F-FOS (in particular 1-kestose) with minor contribution of neoFOS and negligible of levan-type FOS., Projects BIO2010-20508-C04-01, BIO2010-20508-C04-04, BIO2013-48779-C4-1-R and BIO2013-48779-C4-4-R from Spanish Ministry of Economy and Competitiveness supported this research. We thank Lifelong Learning Programme/Erasmus student placement 2012/2013, and Fundacion Ramon Areces for the institutional Grant to the Centro de Biología Molecular Severo Ochoa.

The extracellular β-fructofuranosidase Xd-INV from the yeast Xanthophyllomyces dendrorhous mainly synthesizes the neo-fructooligosaccharides (neo-FOS) neokestose and neonystose. This enzyme is a glycoprotein with a content of 59-67% N-linked carbohydrates and an estimated molecular mass of 160-200. kDa. The extent level of glycosylation affects the thermal behaviour of the enzyme but not its hydrolase and transferase activities, which are optimal at 60-70 °C. The neo-FOS yield of this enzyme was increased from 40 to 168. g/L when the sucrose concentration increased from 420 to 600. g/L and when the reaction was carried out at 60 °C. The neo-FOS levels obtained (168. g/L) in this work are the largest reported for any microbial β-fructofuranosidase. © 2012 Elsevier Ltd.