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AssigneeMount Sinai School of Medicine of the City University of New York
InventorsRobert J. Desnick, David F. Bishop, Yiannis A. Ioannou
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The present invention involves the production of large quantities of human .alpha.-Gal A by cloning and expressing the .alpha.-Gal A coding sequence in eukaryotic host cell expression systems. The eukaryotic expression systems, and in particular the mammalian host cell expression system described herein provide for the appropriate cotranslational and posttranslational modifications required for proper processing, e.g., glycosylation, phosphorylation, etc. and sorting of the expression product so that an active enzyme is produced. In addition, the expression of fusion proteins which simplify purification is described. Using the methods described herein, the recombinant .alpha.-Gal A is secreted by the engineered host cells so that it is recovered from the culture medium in good yield. The .alpha.-Gal A produced in accordance with the invention may be used, but is not limited to, in the treatment in Fabry Disease; for the hydrolysis of .alpha.-galactosyl residues in glycoconjugates; and/or for the conversion of the blood group B antigen on erythrocytes to the blood group O antigen.
A method of producing human .alpha.-galactosidase A, comprising:
A method of producing human .alpha.-galactosidase A, comprising:
A recombinant vector comprising a nucleotide sequence encoding a cleavage site sandwiched between an .alpha.-galactosidase A coding sequence and a protein A domain E coding sequence arranged in the same translational reading frame, controlled by a second nucleotide sequence that regulates the expression of a fusion protein in a mammalian host cell.
A recombinant vector pAGA-pA as deposited with the Agricultural Research Culture Collection having accession number B 18723.
A recombinant vector comprising a nucleotide sequence encoding a cleavage site sandwiched between an .alpha.-galactosidase A coding sequence and a protein antigen coding sequence arranged in the same translational reading frame, controlled by a second nucleotide sequence that regulates the expression of a fusion protein in a mammalian host cell.
Dependencies are inferred from the claim text.