Precise Structure and Anticoagulant Activity of Fucosylated Glycosaminoglycan from Apostichopus japonicus: Analysis of Its Depolymerized Fragments

Abstract

<i>Apostichopus japonicus</i> is one of the most economically important species in sea cucumber aquaculture in China. Fucosylated glycosaminoglycan from <i>A. japonicus</i> (AjFG) has shown multiple pharmacological activities. However, results from studies on the structure of AjFG are still controversial. In this study, the deaminative depolymerization method that is glycosidic bond-selective was used to prepare the depolymerized products from AjFG (dAjFG), and then a series of purified oligosaccharide fragments such as tri-, hexa-, nona-, and dodecasaccharides were obtained from dAjFG by gel permeation chromatography. The 1D/2D NMR and ESI-MS spectrometry analyses showed that these oligosaccharides had the structural formula of l-FucS-α1,3-d-GlcA-β1,3-{d-GalNAc_4S6S-β1,4-[l-FucS-α1,3-]d-GlcA-β1,3-}<i>_n</i>-d-anTal-diol_4S6S (<i>n</i> = 0, 1, 2, 3; FucS represents Fuc_2S4S, Fuc_3S4S, or Fuc_4S). Thus, the unambiguous structure of native AjFG can be rationally deduced: it had the backbone of {-4-d-GlcA-β1,3-d-GalNAc_4S6S-β1-}<i>_n</i>, which is similar to chondroitin sulfate E, and each d-GlcA residue in the backbone was branched with a l-FucS monosaccharide at <i>O</i>-3. Bioactivity assays confirmed that dAjFG and nonasaccharides and dodecasaccharides from AjFG had potent anticoagulant activity by intrinsic FXase inhibition while avoiding side effects such as FXII activation and platelet aggregation.