The Structure-Activity Relationship Of Ginseng Pectin Interacting With Galectin-3

Galectin-3is the only chimeric member of galectin family, composed of the C-terminal carbohydrate-recognition domain (CRD) and N-terminal peptide. Galectin-3can recognize and bind with P-galactosides through the evolutionary conserved CRD. As a cancer-associated protein, Galectin-3plays various roles in the development, metastasis and immune escape of carcinoma cells. Hence, it becomes one of important targets in cancer therapy. Pectin has been shown to inhibit the actions of Galectin-3, which has emerged as an important resource for developing high affinity Galectin-3inhibitors with low toxicity and high water solubility. Modified citrus pectin (MCP) has the best activity among these pectins and most studied. However, the structural features of pectin involved in this activity remain unclear. Pectins have complex structures which are composed of mainly galacturonic acid and an admixture of more than ten kinds of other monosaccharides including galactose. We already studied the isolation and purification of ginseng pectin in our laboratory. The present study aims to further investigate the activities and structure-activity relationship of ginseng pectin on Galectin-3inhibition, which can provide the theoretical foundation for the investigation of Galectin-3inhibitors, pectic drug development and cancer therapy.We firstly studied the activities of pectins with different structures on Galectin-3inhibition. Lactose and MCP were used as the positive controls. Then, we did qualitative and quantitative detection on the binding of RG-I-4to Galectin-3. Finally, we investigated their structure-activity relationships.(1) RG-I-4, extracted from ginseng pectin with classical RG-I domain, had strong inhibitory activity on Galectin-3mediated hemagglutination. At the cellular level, RG-I-4effectively inhibited Galectin-3-mediated cancer cell adhesion, cancer cell homotypic aggregation, endothelial cell migration and the binding of Galectin-3to T-cells, which implied that RG-I-4was an effective Galectin-3inhibitor.(2) RG-I-4, compared to MCP has stronger affinity between and Galectin-3. The affinity was CRD-dependent and could be inhibited completely by lactose.(3) Structure-activity analysis revealed that a) The arabinan side chains positively or negatively regulated the activity depending on their location within the RG-I-4molecule. The galactose residues played pivotal roles on the activity of RG-I-4. b) The activity of galactan chain was proportional to its length up to4Gal residues and largely unchanged thereafter. c) The majority of galactan side chains in RG-I-4were short with low activities. d) The high activity of RG-I-4resulted from cooperation action of these side chains, e) The backbone of the molecule was very important to RG-I-4activity, possibly by maintaining a structural conformation of the whole molecule.f) The isolated backbone could bind with Galectin-3in a CRD-independent manner, which was insensitive to lactose treatment.In summary, we found one RG-I-rich pectin fragment, RG-I-4, which could effectively inhibit Galectin-3mediated biological activities involved in cancer progression. The strong binding activity of RG-I-4to Galectin-3was associated with the cooperation of the side chains and backbone. All the results were valuble for producing highly active pectin-based Galectin-3inhibitors, and they are significant for the drug exploitation of plant pectin.