title: Quantitative Mass-Spectrometric Sequencing of Chitosan Oligomers Revealing Cleavage Sites of Chitosan Hydrolases.
abstract: Partially acetylated chito-oligosaccharides (paCOS) have diverse bioactivities that turn them into promising compounds especially for medical and agricultural applications. These properties likely arise from different acetylation patterns, but determining the sequences of paCOS and producing paCOS with patterns of interest have proven difficult. We present a novel method for sequencing submicrogram amounts of paCOS using quantitative mass spectrometry, allowing one to rapidly analyze the substrate specificities of chitosan hydrolases that can be used to produce paCOS. The method involves four major steps: (i) acetylation of free amino groups in paCOS using a deuterated reagent; (ii) labeling the reducing end with an18O-tag; (iii) quantifying paCOS using [13C2,2H3]-labeled isotopologs as internal standards; (iv) sequencing paCOS by tandem MS. Eventually, this method will aid in developing enzymes with cleavage patterns optimized for producing paCOS with defined patterns of acetylation and specific bioactivities.
authors.affiliation: Institute for Biology and Biotechnology of Plants, University of Münster , Schlossplatz 8, 48143 Münster, Germany.
authors.forename: Stefan
authors.initials: S
authors.lastname: Cord-Landwehr
authors.affiliation: Institute for Biology and Biotechnology of Plants, University of Münster , Schlossplatz 8, 48143 Münster, Germany.
authors.forename: Phillip
authors.initials: P
authors.lastname: Ihmor
authors.affiliation: Institute for Biology and Biotechnology of Plants, University of Münster , Schlossplatz 8, 48143 Münster, Germany.
authors.forename: Anna
authors.initials: A
authors.lastname: Niehues
authors.affiliation: Institute for Organic Chemistry, University of Münster , Corrensstraße 40, 48149 Münster, Germany.
authors.forename: Heinrich
authors.initials: H
authors.lastname: Luftmann
authors.affiliation: Institute for Biology and Biotechnology of Plants, University of Münster , Schlossplatz 8, 48143 Münster, Germany.
authors.forename: Bruno M
authors.identifier: 0000-0001-6067-3205
authors.initials: BM
authors.lastname: Moerschbacher
authors.affiliation: Institute for Hygiene, University of Münster , Robert-Koch-Str. 41, 48149 Münster, Germany.
authors.forename: Michael
authors.initials: M
authors.lastname: Mormann