Epsigam fusion protein

Abstract

Epsi-gam provides a novel fusion protein with the ability to cross-link either of the FcεRI or FcεRII cell surface receptors with an FcγRIIb cell surface receptor in order to block IgE-mediated biological responses.

What is claimed is: 
     
         1 . A method for manufacturing a fusion protein, comprising
 synthesizing a nucleotide sequence comprising of a first restriction site at a first end of the nucleotide sequence and a second restriction site at a second end of the nucleotide sequence;   inserting the nucleotide sequence into a cloning vector;   excising the nucleotide sequence from the cloning vector using a first restriction enzyme corresponding to said first restriction site and a second restriction enzyme corresponding to the second restriction site;   inserting the nucleotide sequence into an expression vector, wherein said expression vector links the expression of an exogenous protein to the glutamine synthetase gene;   transfecting cells with the expression vector using electroporation, wherein the cells are deficient in glutamine synthetase;   incubating the transfected cells; and   collecting a secreted protein from the cells comprising said fusion protein.   
     
     
         2 . The method of  claim 1 , wherein the nucleotide sequence comprises the sequence of SEQ ID: 1. 
     
     
         3 . The method of  claim 2 , wherein the nucleotide sequence is synthesized using gene synthesis techniques. 
     
     
         4 . The method of  claim 1 , wherein the first restriction site and the second restriction site are restricted by the same restriction enzyme. 
     
     
         5 . The method of  claim 1 , wherein the first restriction site comprises a Hind III site, and the second restriction site comprises an EcoRI site. 
     
     
         6 . The method of  claim 1 , wherein the first restriction enzyme comprises HindIII and the second restriction enzyme comprises EcoRI. 
     
     
         7 . The method of  claim 1 , wherein the cloning vector comprises pUC57. 
     
     
         8 . The method of  claim 1 , wherein the cells comprise Chinese Hamster ovary cells. 
     
     
         9 . The method of  claim 1 , wherein electroporation comprises delivering a single pulse at about 300V and 900 uF with resistance set to infinity. 
     
     
         10 . The method of  claim 1 , wherein the cells are incubated at 5% CO2 in a 37 C incubator. 
     
     
         11 . The method of  claim 1 , wherein the fusion protein comprises the sequence of SEQ ID: 2.