Patent ID: 6140059
Filing Date: 2000-10-31
Classification: A61K,A61P,C07K,C12N,Y02A

Abstract:
A method for the production of recombinant truncated HIV envelope glycoprotein gp140 domains that retain their native oligomeric and glycosylated form comprising the following:(i) providing a nucleotide sequence encoding an amino acid sequence comprising a protease recognition site;(ii) providing a nucleotide sequence encoding for the HIV envelope glycoprotein precursor gp160, wherein said sequence lacks a functional wildtype gp120/gp41 proteolytic cleavage site;(iii) inserting the nucleotide sequence of step (i) into the nucleotide sequence of step (ii) at a region corresponding to amino acids 645 to 673 of the transmembrane area of gp41, wherein said numbering scheme is based upon isolate HXB2, thereby creating a mutant HIV envelope gene;(iv) cloning the mutant HIV envelope gene into a suitable expression vector and expressing said mutant HIV envelope gene in eukaryotic cell line to produce a recombinant mutant HIV envelope glycoprotein gp160 comprising said protease recognition site;(v) subjecting the expression product of step (iv) to enzymatic digestion with a suitable protease thereby creating HIV gp140 envelope glycoprotein domains that retain their native oligomeric and glycosylated form; and(vi) isolating and purifying said HIV gp140 from the digestion mixture.