Klotho b, a divergent structural member of the glycosidase I superfamily, is expressed primarily in the liver and pancreas, with lower expression in adipose tissue.1 Like Klotho, Klotho b facilitates binding between FGF19 subfamily members and their receptors via formation of a ternary complex.2 The Klotho b mediated interaction of FGF15 (human FGF19) with FGF Receptor 4 in the liver negatively regulates bile acid synthesis by controlling the secretion of two key bile acid synthase genes, cholesterol 7-a hydroxylase (Cyp7a1) and sterol 12-a hydroxylase (Cyp8b1).2-4 Klotho b is also a cofactor for the interaction of FGF21 with FGF Receptor 1c in adipocytes, which allows FGF21 to stimulate GLUT1 expression, upregulating adipocyte insulin-dependent glucose uptake.2-3, 5 The 1043 amino acid (aa) type I transmembrane protein is composed of a 51 aa signal sequence, a 943 aa extracellular domain (ECD) containing two glycosidase-like regions, a 21 aa transmembrane domain, and 28 aa intracellular tail. Since Klotho-related proteins lack critical active site Glu residues present in b-glycosidases, it was initially unclear whether they were functional enzymes.1, 6 However, glucuronidase activity has since been demonstrated for Klotho, indicating that physiologically relevant enzymatic activity for Klotho b is also possible.7 The extracellular domain shares 79%, 79%, 80% and 67% identity with human, bovine, canine and rat Klotho b, respectively. The low identity with rat reflects aa discordance within rodent ECD.
A DNA sequence encoding the extracellular domain of mouse Klotho Beta was fused to a polyhistdine tag at the C-terminus. The protein was expressed in a mouse myeloma cell line, NS0.
Molecular Mass: 110.2kD. As a result of glycosylation, the recombinant mouse Klotho Beta migrates as an approximately 140kD protein in SDS-PAGE under reducing conditions.
Biological Activity: Measured by its ability to stimulate proliferation of NIH 3T3 cells. The ED50 for this effect is typically 0.75-3ug/ml in the presence of 2ug/ml of rhFGF-21
Endotoxin: (same/less than) 1EU/ug of the protein as determined by the LAL method.
Storage and Stability:
May be stored at 4 degrees C for short-term only. For long-term storage, aliquot and store at -20 degrees C. Aliquots are stable for at least 6 months at -20 degrees C. For maximum recovery of product, centrifuge the original vial after thawing and prior to removing the cap. Further dilutions can be made in assay buffer.