Biologically active Eph family ligands

Abstract

A novel ligand (Efl-6) that binds the Elk subfamily of Eph receptors is identified, and methods for making the soluble Elf-6 ligand in biologically active form is described. A cDNA clone encoding this novel protein enables production of the recombinant protein, which is useful to support neuronal and other eph receptor-bearing cell populations.

1 . An isolated and purified nucleic acid molecule encoding Efl-6 protein wherein the sequence of said nucleic acid is selected from the group consisting of: 
 (a) the sequence of the DNA encoding mature Efl-6 protein contained in the plasmid pBluescriptSK 14  Efl-6 as deposited with the American Type Culture Collection on Oct. 19, 1995 and designated as 97319;    (b) the sequence of the DNA encoding mature Efl-6 protein as set forth in FIG. 1;    (c) DNA sequences that hybridize under moderately stringent conditions to the DNA of (a) or (b) and which encode a protein that binds a receptor belonging to the Elk subfamily of Eph receptors; and    (d) DNA sequences that are degenerate as a result of the genetic code to a DNA sequence of (a), (b), or (c) and which encode an Efl-6 protein that binds a member of the Elk subclass of Eph receptors.    
     
     
         2 . Isolated and purified mature Efl-6 protein having an amino acid sequence as set forth in FIG. 1.  
     
     
         3 . An isolated nucleic acid encoding the extracellular domain of Efl-6 (sEfl-6) having a sequence selected from the following: 
 (a) the sequence set forth from about nucleotide 274 to about nucleotide 873 of FIG. 1; and    (b) a sequence which encodes the extracellular domain of Efl-6 as set forth in FIG. 1.    
     
     
         4 . Purified sEfl-6 encoded by the nucleotide sequence of  claim 3 .  
     
     
         5 . (sEfl-6)n comprising the sEfl-6 protein according to  claim 4 , wherein n is 2 or greater.  
     
     
         6 . Efl-6 ligandbody comprising soluble Efl-6 protein according to  claim 4  and the Fc portion of IgG.  
     
     
         7 . A vector which comprises a nucleic acid molecule of  claim 1 .  
     
     
         8 . A vector according to  claim 7  wherein the nucleic acid molecule is operatively linked to an expression control sequence capable of directing its expression in a host cell.  
     
     
         9 . A host cell containing a vector according to  claim 8 .  
     
     
         10 . A vector which comprises a nucleic acid molecule of  claim 3 .  
     
     
         11 . A vector according to  claim 10  wherein the nucleic acid molecule is operatively linked to an expression control sequence capable of directing its expression in a host cell.  
     
     
         12 . A host cell containing a vector according to  claim 11 .  
     
     
         13 . A method of producing Efl-6 ligand which comprises growing cells of a host according to  claim 8  under conditions permitting production of the ligand, and recovering the ligand so produced.  
     
     
         14 . A method of producing Efl-6 soluble ligand which comprises growing cells of a host according to  claim 11  under conditions permitting production of the ligand, and recovering the ligand so produced.  
     
     
         15 . An antibody which specifically binds the ligand of  claim 2  or  4 .  
     
     
         16 . An antibody according to  claim 15  which is a monoclonal antibody.