Structural Elucidation of Enzymatically Synthesized Galacto-oligosaccharides Using Ion-Mobility Spectrometry-Tandem Mass Spectrometry.
Galacto-oligosaccharides (GOS) represent a diverse group of well-characterized prebiotic ingredients derived from lactose in a reaction catalyzed with β-galactosidases. Enzymatic transgalactosylation results in a mixture of compounds of various degrees of polymerization and types of linkages. Because structure plays an important role in terms of prebiotic activity, it is of crucial importance to provide an insight into the mechanism of transgalactosylation reaction and occurrence of different types of β-linkages during GOS synthesis. Our study proved that a novel one-step method, based on ion-mobility spectrometry-tandem mass spectrometry (IMS-MS/MS), enables complete elucidation of GOS structure. It has been shown that β-galactosidase from Aspergillus oryzae has the highest affinity toward formation of β-(1→3) or β-(1→6) linkages. Additionally, it was observed that the occurrence of different linkages varies during the reaction course, indicating that tailoring favorable GOS structures with improved prebiotic activity can be achieved by adequate control of enzymatic synthesis.
Department of Biochemical Engineering and Biotechnology, Faculty of Technology and Metallurgy, University of Belgrade , Karnegijeva 4, 11000 Belgrade, Serbia.
Milica
M
Carević
Department of Biochemical Engineering and Biotechnology, Faculty of Technology and Metallurgy, University of Belgrade , Karnegijeva 4, 11000 Belgrade, Serbia.
Dejan
D
Bezbradica
Department of Biochemical Engineering and Biotechnology, Faculty of Technology and Metallurgy, University of Belgrade , Karnegijeva 4, 11000 Belgrade, Serbia.
Katarina
K
Banjanac
Department of Biochemical Engineering and Biotechnology, Faculty of Technology and Metallurgy, University of Belgrade , Karnegijeva 4, 11000 Belgrade, Serbia.
Ana
A
Milivojević
INRA , UR1268 Biopolymers Interactions Assemblies, F-44316 Nantes, France.
Mathieu
M
Fanuel
INRA , UR1268 Biopolymers Interactions Assemblies, F-44316 Nantes, France.
Hélène
H
Rogniaux
INRA , UR1268 Biopolymers Interactions Assemblies, F-44316 Nantes, France.
David
D
Ropartz
Faculty of Chemistry, University of Belgrade , Studentski trg 12, 11000 Belgrade, Serbia.
Dušan
D
Veličković
D001236:Aspergillus oryzae / Q000201:enzymology*; D055162:Biocatalysis; D005656:Fungal Proteins / Q000737:chemistry* / Q000378:metabolism; D005690:Galactose / Q000737:chemistry / Q000378:metabolism; D015394:Molecular Structure; D009844:Oligosaccharides / Q000737:chemistry* / Q000378:metabolism; D053719:Tandem Mass Spectrometry / Q000295:instrumentation / Q000379:methods*; D001616:beta-Galactosidase / Q000737:chemistry* / Q000378:metabolism
D005656:Fungal Proteins; D009844:Oligosaccharides; D001616:beta-Galactosidase; D005690:Galactose
A. oryzae; IMS-MS/MS; galacto-oligosaccharides; mass spectrometry; structure