The University of British Columbia

Richard A Warren

Douglas G Kilburn

Neil R Gilkes

Robert C Miller, Jr

Robert C Miller

Novel polypeptide compositions and methods for their use are provided comprising fusion proteins in which the polysaccharide binding domain or functional portion thereof of a polysaccharidase is fused to a heterologous protein or is conjugated to a chemical moiety. The compositions can be synthesized or prepared by recombinant DNA technology. The compositions find use as removable labels.

Polysaccharide binding fusion proteins and conjugates

Fusion proteins or conjugates are provided containing an amino acid sequence having a substrate binding region of a polysaccharidase such as cellulase that binds to a .beta.-1,4-glycan matrix such as cellulose. The substrate binding region is essentially without polysaccharidase activity. In the fusion protein, the substrate binding region is fused or chemically linked to a polypeptide such as an enzyme, a hormone, an immunoglobulin or a protein dye. By contacting the fusion protein with a .beta.-1,4-glycan matrix, the substrate binding region binds to the matrix to immobilize the polypeptide on the matrix. The polypeptide or fusion protein can be removed from the matrix with a protease acting on a protease recognition sequence or with a solution having a low ionic strength or high pH. In the conjugate, the substrate binding region is joined such as by covalent bonding to a non-protein chemical moiety such as a dye, chromophore, fluorescor, radionuclide or enzyme co-factor. By contacting the conjugate with a .beta.-1,4-glycan matrix, the substrate binding region binds to the matrix to immobilize the chemical moiety on the matrix. The conjugate or chemical moiety can be removed with a protease acting on a protease recognition sequence or with a solution having a low ionic strength or high pH.

A fusion protein is prepared containing a polypeptide such as an enzyme and an amino acid sequence having a substrate binding region of a polysaccharidase such as cellulase that has essentially no polysaccharidase activity. By contacting the fusion protein with an affinity matrix containing a substrate such as cellulose for the cellulase substrate binding region, the substrate binding region binds to the affinity matrix to immobilize the polypeptide. The polypeptide can be purified by separating the fusion protein or polypeptide from the affinity matrix. The polypeptide can be separated by cleaving the protein with a Cellulomonas fimi protease.

Cellulose binding fusion proteins having a substrate binding region of

A fusion protein is prepared containing a polypeptide such as an enzyme and an amino acid sequence having a substrate binding region of a polysaccharidase such as cellulase that has essentially no polysaccharidase activity. By contacting the fusion protein with an affinity matrix containing a substrate such as cellulose for the cellulase substrate binding region, the substrate binding region binds to the affinity matrix to immobilize the polypeptide. The polypeptide can be purified by separating the fusion protein or polypeptide from the affinity matrix.

Growing community of inventors

Vancouver, Canada

Richard A Warren