Crystal of Human Glycoprotein VI Collagen Binding Domain

Abstract

A crystal comprising the collagen binding domain of human GPVI is provided and defined by structural atomic coordinates. Employing computer modeling based on the crystal structure, including methods for identifying inhibitors of GPVI collagen binding activity, methods for screening libraries of compounds for potential to bind to the GPVI collagen binding domain, and methods of identifying a compound useful for the treatment of a GPVI-mediated disorder, are also provided.

1 . A method of identifying an inhibitor of GPVI collagen binding activity, the method comprising: (1) designing or selecting a potential inhibitor using a three-dimensional structure of the GPVI collagen binding domain comprising main chain and side chain atoms and as defined by atomic coordinates set forth in Table 2, plus or minus a root mean square deviation for the main chain atoms of not greater than 2 Angstroms; (2) synthesizing or obtaining the potential inhibitor; and (3) determining whether the potential inhibitor inhibits the activity of GPVI, wherein designing or selecting a potential inhibitor employs computer modeling. 
     
     
         2 . The method according to  claim 1 , wherein determining whether the potential inhibitor inhibits the activity of GPVI is performed using a protein binding assay or an inhibition assay. 
     
     
         3 . The method according to  claim 1 , wherein determining whether the potential inhibitor inhibits the activity of GPVI is performed using a cellular-based assay. 
     
     
         4 . The method according to  claim 1 , wherein determining whether the potential inhibitor inhibits the activity of GPVI comprises contacting the potential inhibitor with GPVI in the presence of a known ligand, determining the ability of the potential inhibitor to compete with the known GPVI collagen binding domain ligand and screening the potential inhibitor of GPVI in an assay for efficacy in inhibiting GPVI. 
     
     
         5 . The method according to  claim 4 , wherein the known ligand is selected from the group consisting of collagen, CRP and convulxin. 
     
     
         6 . A method for designing binding agents that interact with the GPVI collagen binding domain, the method comprising: employing at least some of the atomic coordinates set forth in Table 2. 
     
     
         7 . A method of screening a library of compounds for potential to bind to the GPVI collagen binding domain, the method comprising: using at least some of the atomic coordinates set forth Table 2 to perform computational docking of the library of compounds to the GPVI collagen binding domain. 
     
     
         8 . A method of identifying a compound useful for the treatment of a GPVI-mediated disorder, comprising the steps of: (a) using a three-dimensional structure of the GPVI collagen binding domain as defined by at least some of the atomic coordinates set forth in Table 2, and (b) employing the structure to design, modify, or select a compound that modulates GPVI activity. 
     
     
         9 . The method according to  claim 8 , wherein the GPVI modulator is useful in the prevention or treatment of a vascular disorder. 
     
     
         10 . The method according to  claim 9 , wherein the vascular disorder comprises a thrombotic disorder. 
     
     
         11 . The method according to  claim 8 , wherein the GPVI collagen binding domain comprises at least some or all of residues K41, S43, S44, R46, Q48, Q50, L53, F54, I55, P56, K59, S61, L62, Y66, and R166. 
     
     
         12 . A method of designing or selecting agents that bind to the collagen binding domain of GPVI according to  claim 2 , the method comprising the use of at least some of the atomic coordinates set forth in Table 2 to perform computer modeling. 
     
     
         13 . The method according to  claim 12 , wherein two designed or selected agents are connected by a crosslinker of appropriate length to permit bivalent binding to the dimeric form of GPVI observed in the crystal.