| Wnt signaling is involved in variety of developmental processes including cell fate determination, cell polarity, tissue patterning and control of cell proliferation. Members of the Frizzled family of proteins serve as receptors for the Wnt signaling pathway. The founding member of this family was identified in Drosophila based on its role in tissue polarity in the adult cuticle and named for the disorganized appearance of bristle hairs on the mutant. Ten mammalian 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, a variable-length linker region, a seven-pass transmembrane domain, and a variablelength C-terminal tail. One of the most conserved regions of the Frizzled (Frz) proteins is the extracellular cysteine-rich domain (CRD) which spans approximately 120aa and contains 10 invariant cysteines (1). Human Frz-5 shows 95% amino acid identity to mouse Frz-5 in the CRD region. In the mouse, Frz-5 is expressed in adult tissues (heart and kidney), as well as embryonic tissues (telencephalon, eye, and lung bud) (2,3). Null mutations in Frz-5 reveal that it plays a role in the formation and maintenance of the extra-embryonic vasculature (3). Functional interactions with Frz-5 have been reported for Wnt5a, Wnt10b, Wnt2b, and Wnt 7a, implicating these Wnts as ligands for the Frz-5 receptor (3-5). Two distinct Wnt signal transduction pathways have been characterized. One is the canonical Wnt/beta-catenin pathway that is involved in diverse biological mechanisms such as dorsal/ventral development in Xenopus embryos and mammalian tumor formation. Frz-5 is implicated in this pathway based on its ability to induce beta-catenin target genes in the presence of ligand (5). However, Frz-5 is also implicated in beta-catenin independent pathways (4,6). Since the CRD of hFrz-5, like mFrz-8, binds cell-surface Xenopus Wnt-8 (Xwnt-8) protein 7, this product is tested to antagonize Wnt signaling for inhibition of secondary axes induced by Xwnt-8 mRNA. Source: N- Human CD33 Signal Peptide (Met1-Ala16)- Human Frizzled-5 (Ala27-Pro167)- IEGRMD- Human IgG1 (Pro100-Lys330)- C A DNA sequence encoding the signal peptide from human CD33 joined with the extracellular cysteine-rich domain (CRD) of human Frizzled-5 (Ala27-Pro167) (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 human Frizzled-5/Fc is a disulfide-linked homodimeric protein. Based on N-terminal amino acid sequencing, the recombinant human Frizzled-5/Fc starts at Ala27. Each monomer consists of 378 amino acids and has a calculated molecular mass of 42.5kD. As a result of glycosylation, the recombinant monomer migrates as an approximately 55-60kD protein in SDS-PAGE under reducing conditions. Endotoxin Level: 0.01EU/1ug (LAL). Activity: Measured by its ability to interfere with the axis-inducing activity of Xwnt-8 mRNA in early Xenopus embryos. In a population of embryos, addition of 1ng of human Frizzled-5/Fc protein per embryo resulted in 50-100% reduction in the number of secondary axes produced by 10-20pg of Xwnt-8 mRNA. Reconstitution: It is recommended that sterile PBS be added to the vial to prepare a working stock solution of no less than 50ug/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 |
