UP-HILIC-MS/MS to Determine the Action Pattern of Penicillium sp. Dextranase

作者：Yi, Lin; Sun, Xue; Du, Kenze; Ouyang, Yilan; Wu, Chengling; Xu, Naiyu^*; Linhardt, Robert J.; Zhang, Zhenqing

来源：Journal of the American Society for Mass Spectrometry, 2015, 26(7): 1174-1185.

DOI：10.1007/s13361-015-1117-y

• HILIC
• MS/MS
• Dextran
• Oligosaccharide
• Sequence
• Dextranase
• Action pattern

Investigation of the action pattern of enzymes acting on carbohydrates is challenging, as both the substrate and the digestion products are complex mixtures. Dextran and its enzyme-derived oligosaccharides are widely used for many industrial applications. In this work, a new method relying on ultra-performance hydrophilic interaction liquid chromatography quadrupole time-of-flight tandem mass spectrometry (UP-HILIC- Q/TOF-MS/MS) was developed to analyze a complex mixture of dextran oligosaccharide products to determine the action pattern of dextranase. No derivatization of oligosaccharides was required and the impact of the alpha- and beta-configurations of the native oligosaccharides on the chromatographic separation was eliminated. The 1 -> 6, 1 -> 3, 1 -> 4 backbone linkages and the branch linkages of these oligosaccharides were all distinguished from diagnostic ions in their MS/MS spectra, including fragments corresponding to (0,2)A, (0,3)A, (0,4)A, B-H2O, (2,5)A, and (3,5)A. The sequences of the oligosaccharide products were similarly established. Thus, the complex oligosaccharide mixtures in dextran digestion products were profiled and identified using this method. The more enzyme-resistant structures in dextran were established using much less sample, labor, time, and uncertainty than in previous studies. This method provides an efficient, sensitive, and straightforward way to monitor the entire process of digestion, establish the action pattern of the dextranase from Penicillium sp., and to support the proper industrial application of dextranase.

• 出版日期2015-7
• 单位苏州大学