The Wnt genes encode a large family of glycoproteins that are essential in development and tissue maintenance (1). Members of the Frizzled family of proteins serve as receptors for the Wnt signaling pathway (2). Ten mouse and human Frizzled genes have been identified to date. The predicted structure of Frizzled proteins is similar among all family members, containing a divergent N-terminal signal peptide, a highly conserved extracellular cysteine-rich domain (CRD), a variable-length linker region, a sevenpass transmembrane region, and a variable-length C-terminal cytoplasmic domain. The CRD, which comprises the Wnt binding site, spans about 130 amino acid residues and contains ten invariant cysteine residues. Mouse Frizzled-7 contains 572 amino acid residues and shares 99% identity with the human orthologue in the CRD (3,4). Frizzled-7 mRNA has been detected in relatively large amounts in adult skeletal muscle, plancenta and heart, and fetal kidney and lung (3). Several Frizzled-dependent signaling pathways exist (2). Their activation depends on the Wnt ligand and the cell context. Frizzled-7 can activate canonical Wnt/beta-Catenin signaling, as well as a non-canonical pathway involved in tissue morphogenesis (4,5). Source: N- Mouse Frizzled-7 (Met1-Leu185)- IEGRMD- Human IgG1 (Pro100-Lys330)- C A DNA sequence encoding amino acid residues Met1-Leu185 of mouse Frizzled-7 (Wang, Y. et al., 1996, J. Biol. Chem. 271: 4468-4476) was fused to the Fc region of human IgG1 via a peptide linker. The chimeric protein was expressed in a mouse myeloma cell line, NS0. Molecular Mass: The recombinant mature mouse Frizzled-7/Fc chimera, generated by proteolytic removal of the signal peptide, is a disulfide linked homodimeric protein. Based on amino acid sequencing, the N-terminus of the recombinant protein was blocked, suggesting that the recombinant protein may start at Gln33 as predicted. The calculated molecular mass of the mature recombinant protein is ~43kD. As a result of glycosylation, the recombinant protein migrates as an ~55-60kD protein in SDS-PAGE under reducing conditions. Endotoxin Level: 1.0EU per 1ug of the cytokine as determined by the LAL method. Activity: Measured by its ability to bind biotinylated Wnt-3a in a functional ELISA. When 100ul of the reaction mixture is captured on a streptavidin-coated plate, 200ng/ml of biotinylated rmWnt-3a will bind rmFrizzled-7/Fc with a linear range of 10-1000ng/ml. Reconstitution: It is recommended that sterile PBS be added to the vial to prepare a working stock solution of no less than 100ug/ml. The carrier-free protein should be used immediately upon reconstitution to avoid losses in activity due to non-specific binding to the inside surface of the vial. For long term storage as a dilute solution, a carrier protein (e.g. 0.1% HSA or BSA) should be added to the vial. Storage: Lyophilized samples are stable for up to twelve months from date of receipt at -20C to -70C. Upon reconstitution, this cytokine, in the presence of a carrier protein, can be stored under sterile conditions at 2-8C for one month or at -20C to -70C in a manual defrost freezer for three months without detectable loss of activity. Avoid repeated freeze-thaw cycles.
