Production and purification of a protein fused to a binding prot

Details Production and purification of a protein fused to a binding prot Production and purification of a protein fused to a binding prot

1995-01-12

1997-07-01

Guzo, David

Chemistry: molecular biology and microbiology

Micro-organism, tissue cell culture or enzyme using process...

Recombinant dna technique included in method of making a...

4353201, C12P 2100, C12N 1570

Patent

active

056437587

ABSTRACT:
Methods and products are provided for producing and/or purifying virtually any hybrid polypeptide molecule employing recombinant DNA techniques. More specifically, a DNA fragment coding for a protein molecule, e.g. a polypeptide or portion thereof, is fused to a DNA fragment coding for a binding protein, such as the gene coding for the maltose binding protein. The fused DNA is inserted into a cloning vector and an appropriate host transformed. Upon expression, a hybrid polypeptide is produced which can be purified by contacting the hybrid polypeptide with a ligand or substrate to which the binding protein has specific affinity, e.g. by affinity chromatography. The hybrid polypeptide so purified may in certain instances be useful in its hybrid form, or it may be cleaved to obtain the protein molecule itself by, for example, linking the DNA fragments coding for the target and binding proteins with a DNA segment which codes for a peptide which is recognized and cut by a proteolytic enzyme, such as Factor Xa. The present invention also relates to certain vectors useful in practicing the above process.

REFERENCES:
Rasmussen et al. (1984) J. Bacteriol. 160:612-617.
Ito et al. (1981) Cell 24:707-717.
Nagai et al. (1984) Nature 309:810-812.
Duplay et al. (1984) J. Bio. Chem. 259:10606-10613.
Bassford et al. (1979) 139:19-31.

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