| Vitronectin is a large glycoprotein found in blood and the extracellular matrix (ECM). The gene for vitronectin encodes a 19 amino acid (aa) signal peptide and a 459 aa protein. The amino terminal 130 aa residues of vitronectin contains multiple binding sites for a variety of structures. Included is a site for binding to plasminogen activator inhibitor-1 (PAI-1) and urokinase receptor, an (RGD) sequence that binds avb3, avb5, aavb1, aIIbb3, avb6, and avb8 integrins, a stretch of acidic amino acids that includes two sulfated tyrosine residues that bind thrombin-anti-thrombin III complexes, and a collagen binding site. The major part of the vitronectin molecule (aa 132-459) contains six hemopexin-like repeats. The carboxyl-terminal end of vitronectin also has multiple sites and functions. It contains a stretch of basic amino acids that binds the acidic amino acids of the amino-terminal region, thereby stabilizing vitronectins three dimensional structure. The carboxyl-terminal end also contains a plaminogen binding site, a heparin binding site that binds complement factor C7, C8 or C9, a glycosaminoglycan binding site, and a second PAI-1 binding site (aa 348-370). Vitronectin also contains an endogenous cleavage site, plus cleavage sites for elastase, thrombin and plasmin. Vitronectin has also been shown to bind IGF-2 and TGF-b. The apparent molecular weight of human vitronectin is 75 kDa, with ~30% of its molecular mass being attributed to glycosylation at 3 different sites. In blood and plasma, vitronectin is found predominantly as a single chain monomer. It can also be found as a dimer after endogenous cleavage. The dimer is composed of a 65kD and 10kD component held together by a disulfide bond. Binding of thrombin-anti-thrombin II complex or complement leads to an unfolding of vitronectin. Unfolding of vitronectin generates disulfide-linked multimers that are found in platelet secretions and extracellular matrix. Vitronectin is produced at high levels by the liver and many tumors. As might be expected by its structure, Vitronectin is involved in a number of biological activities including cell adhesion, cell spreading and migration, cell proliferation, extracellular anchoring, fibrinolysis, hemostasis, and complement mediated immune defense. Source: A DNA sequence encoding human Vitronectin (Met 1-Leu 478) (Suzuki, S. et al., 1985, EMBO J. 4:2519-2524) was expressed in a mouse myeloma cell line, NS0. Molecular Mass: Based on N-terminal sequencing, the recombinant human Vitronectin protein starts at Asp 20 and has a calculated molecular mass of 52.3kD. As a result of glycosylation, the full-length recombinant protein migrates as an approximately 70-80kD band in SDS-PAGE under reducing conditions. Activity: Measured by the ability of the immobilized protein to support the adhesion of DU145 cells. When 5x10e4 cells/well are added to Vitronectin coated plates (5ug/ml and 100ul/well), approximately 55-75% will adhere after 30 minutes at 37C. Storage and Stability: Lyophilized powder may be stored at -20C. Stable for 12 months. Reconstitute with sterile PBS, 01% BSA or HSA. Aliquot and store at -20C/-70C. Reconstituted product is stable for 6 months at --20C/-70C. 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. |
