Porcine alpha2delta-1 calcium channel submit cDNA and soluble secreted alpha2delta-1 subunit polypeptides

Abstract

Soluble α 2 δ-1 subtype polypeptides. Methods for cloning, expression and purification of freely soluble α 2 δ-1 subtype polypeptides.

1 . A purified or isolated nucleic acid encoding a eukaryotic secreted soluble cerebral cortical voltage-dependent calcium channel α 2 δ-1 subunit polypeptide.  
     
     
         2 . A purified or isolated nucleic acid according to  claim 1 , comprising a polynucleotide having at least 90% identity with the sequence encoding from amino-acid 1 to between amino-acids 1009 to 1083 of SEQ ID NO:5 or SEQ ID NO:14.  
     
     
         3 . A purified or isolated nucleic acid according to  claim 1 , having at least 90% identity with the sequence encoding from amino-acid 1 to between amino-acids 1043 and 1088 of SEQ ID NO:5 or SEQ ID NO:14.  
     
     
         4 . A purified or isolated nucleotide sequence according to  claim 1  wherein said sequence is the sequence of SEQ ID NO:2, SEQ ID NO:3, SEQ ID NO:4, SEQ ID NO:19, SEQ ID NO:20, or SEQ ID NO:21.  
     
     
         5 . A purified or isolated nucleic acid, having at least 90% identity with the nucleotide sequence of SEQ ID NO:1.  
     
     
         6 . A purified or isolated polynucleotide comprising at least 10 consecutive nucleotides of the nucleotide sequence of SEQ ID NO:1.  
     
     
         7 . A polynucleotide probe or primer hybridizing, under stringent conditions, with the nucleic acid according to  claim 5 .  
     
     
         8 . A method for the amplification of a nucleic acid encoding a eukaryotic secreted soluble cerebral cortical voltage-dependent calcium channel α 2 δ-1 subunit polypeptide, said method comprising the steps of: 
 (a) contacting a test sample suspected of containing the target secreted soluble α 2 δ-1 subunit nucleic acid, or a sequence complementary thereto, with an amplification reaction reagent comprising a pair of amplification primers located on either side of the α 2 δ-1 subunit nucleic acid region to be amplified, and    (b) optionally, detecting the amplification products.    
     
     
         9 . A kit for the amplification of a nucleic acid encoding a secreted soluble α 2 δ-1 subunit polypeptide, or a complementary sequence thereto in a test sample, wherein said kit comprises: 
 (a) a pair of oligonucleotide primers which can hybridize, under stringent conditions, to the secreted soluble α 2 δ-1 subunit nucleic acid region to be amplified; and    (b) optionally, the reagents necessary for performing the amplification reaction.    
     
     
         10 . A recombinant vector comprising a nucleic acid according to  claim 1 .  
     
     
         11 . A recombinant host cell comprising a nucleic acid according to  claim 1 .  
     
     
         12 . A method for producing a secreted soluble α 2 δ-1 subunit wherein said method comprises the steps of: 
 (a) inserting the nucleic acid encoding the desired α 2 δ-1 subunit polypeptide in an appropriate vector;    (b) culturing, in an appropriate culture medium, a host cell previously transformed or transfected with the recombinant vector of step (a);    (c) harvesting the culture medium thus obtained or lyse the host cell, for example by sonication or osmotic shock;    (d) separating or purifying, from said culture medium, or from the pellet of the resultant host cell lysate, the thus produced α 2 δ-1 subunit polypeptide of interest.    
     
     
         13 - 15 . (canceled).