Chemical O -sulfation of N -sulfoheparosan: a route to rare N -sulfo-3- O -sulfoglucosamine and 2- O -sulfoglucuronic ac
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ORIGINAL ARTICLE
Chemical O-sulfation of N-sulfoheparosan: a route to rare N-sulfo-3-O-sulfoglucosamine and 2-O-sulfoglucuronic acid Lufeng Yan 1,2 & Paul Brodfueher 2 & Li Fu 2 & Fuming Zhang 2 & Shiguo Chen 1 & Jonathan S. Dordick 2 & Robert J. Linhardt 2 Received: 6 May 2020 / Revised: 27 July 2020 / Accepted: 6 August 2020 # Springer Science+Business Media, LLC, part of Springer Nature 2020
Abstract Heparosan, the capsular polysaccharide of E. coli K5 is currently used as the starting material in the chemoenzymatic synthesis of heparan sulfate and the structurally related anticoagulant drug heparin. Base hydrolysis of N-acetyl groups and their subsequent N-sulfonation, are used to prepare N-sulfoheparosan an intermediate of biosynthesis. In the present study, when excess sulfonation reagent was used during N-sulfonation, some O-sulfation also took place in the N-sulfoheparosan product. After a nearly full digestion, a hexasaccharide fraction exhibited resistance to heparin lyase II. Excessive digestion by heparin lyase II and structural identification by NMR and mass spectroscopy indicated that the resistant hexasaccharide fraction has two structures, ΔUAGlcNS-GlcA2S-GlcNS-GlcA-GlcNS and ΔUA-GlcNS-GlcA- GlcNS3S-GlcA-GlcNS in similar amounts. The 2-sulfated structure exhibited partial resistance to heparin lyase II; however the structure of ΔUA-GlcNS-GlcA-GlcNS3S was completely resistant to heparin lyase II. Keywords Chemical O-sulfation . N-sulfoheparosan . 2-O-sulfoglucuronic acid . N-sulfo-3-O-sulfoglucosamine . Resistance to heparin lyase II
Introduction Heparan sulfate and the structurally related heparin are biosynthesized in the Golgi compartments of animal cells [1]. Both of these glycosaminoglycans play critical biological roles in regulating cell growth and differentiation [2], as well as impacting infectious disease [3]. Heparin has an important pharmacological role as a frequently used clinical anticoagulant drug [4]. Heparosan, the capsular polysaccharide of Escherichia coli K5, has the repeating structure [→4) β-D-glucuronic acid (GlcA) (1 → 4) N-acetyl-α-D-glucosamine (GlcNAc) (1→] Electronic supplementary material The online version of this article (https://doi.org/10.1007/s10719-020-09939-7) contains supplementary material, which is available to authorized users. * Robert J. Linhardt [email protected] 1
National-Local Joint Engineering Laboratory of Intelligent Food Technology and Equipment, College of Biosystems Engineering and Food Science, Zhejiang University, Hangzhou 310058, China
2
Center for Biotechnology and Interdisciplinary Studies, Rensselaer Polytechnic Institute, Troy, NY 12180, USA
(Fig. 1a) and is currently used as the starting material in the chemoenzymatic synthesis of heparan sulfate and heparin [5]. N-sulfoheparosan (NSH) has the repeating structure [→4) β-D-GlcA (1 → 4) N-sulfo-α-D-GlcN (1→] (Fig. 1b) and is an early intermediate in the biosynthesis of heparan sulfate and heparin [6]. The heparosan polysaccharide (Fig. 1a) is modified through the action of N-deacetylase-N
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