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

Abstract

Apostichopus japonicus is one of the most economically important species in sea cucumber aquaculture in China. Fucosylated glycosaminoglycan from A. japonicus (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-GalNAc4S6S-β1,4-[l-FucS-α1,3-]d-GlcA-β1,3-}n-d-anTal-diol4S6S (n = 0, 1, 2, 3; FucS represents Fuc2S4S, Fuc3S4S, or Fuc4S). Thus, the unambiguous structure of native AjFG can be rationally deduced: it had the backbone of {-4-d-GlcA-β1,3-d-GalNAc4S6S-β1-}n, which is similar to chondroitin sulfate E, and each d-GlcA residue in the backbone was branched with a l-FucS monosaccharide at O-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.