Abstract
Glycosaminoglycans (GAGs) are polysaccharides with structural and functional diversity. These molecules have been demonstrated to be involved in a wide range of biological activities such as cell adhesion, cell mobility and cell proliferation through interacting with various cellular proteins, such as protease, cytokines, growth factors, adhesion molecules and so on. There are six classes of GAGs, including heparin sulfate (HS), heparin, chondroitin sulfate (CS), dermatan sulfate (DS), keratin sulfate (KS) and hyaluronic acid (HA); of all the GAG family members, heparin is the most well studied and has been used clinically as an anticoagulant since 1935. Recent experiments have shed light on the physiological significance of heparin which is essential for the storage of specific granule proteases in mast cells. The details of the interaction between heparin and positively charged proteins are not known except the electrostatic interactions seem to play an important role in this biomolecular association. With the characterization of more heparin-binding proteins, it was understood that the binding epitopes can also be defined by sequentially remote residues that form an optimal binding surface. However, none of these studies, such as the early studied key structural motifs of heparin-binding sequences XBBXBX and XBBBXXBX, have completely addressed the binding pattern for heparin-protein interactions. Therefore, we use a structure-guided approach, termed “protein grafting”, to elucidate the protein-heparin binding pattern. We targeted the heparin utilizing a thermal stable domain, B1 domain of IgG-binding protein G, and its derivatives because of its firmly packed conformation to accomplish spatial binding information. Through the planer four-beta-sheet scaffold design of B1 domain, the number and position of the positive amino acids within were verified by applying fluorophore-assisted carbohydrate electrophoresis analysis (FACE). The results reveal that the specific spacing and favored residue (arginine) seemed to be critical for heparin binding process.