This Mouse c-MET overexpression lysate was created in HEK293 Cells and intented for use as a Western blot (WB) positive control. Purification of c-MET protein (Cat: 50622-M08H) from the overexpression lysate was verified.
A DNA sequence encoding the mouse MET (NP_032617.2) extracellular domain (Met 1-Asn 929) was fused with a polyhistidine tag at the C-terminus.
The recombinant mouse Met is a heterodimer composed of the proteolytically cleaved α and β subunits. The α and β heterodimer consists of 916 amino acids and has a predicted molecular mass of 102 (α =32 + β=70) kDa. The apparent molecular mass of the rmMET heterodimer thus is approximately 43 kDa and 85-95 kDa respectively in SDS-PAGE under reducing conditions due to glycosylation.
Mouse c-MET HEK293 Overexpression Lysate: 使用指南
Cell lysate was prepared by homogenization of the over-expressed cells in ice-cold modified RIPA Lysis Buffer with cocktail of protease inhibitors (Sigma). Cell debris was removed by centrifugation. Protein concentration was determined by Bradford assay (Bio-Rad protein assay, Microplate Standard assay). The cell lysate was boiled for 5 min in 1 x SDS loading buffer (50 mM Tris-HCl pH 6.8, 12.5% glycerol, 1% sodium dodecylsulfate, 0.01% bromophenol blue) containing 5% b-mercaptoethanol, and lyophilized.
Hepatocyte growth factor receptor (HGFR), also known as c-Met or mesenchymal-epithelial transition factor (MET), is a receptor tyrosine kinase (RTK) that is overexpressed and/or mutated in a variety of malignancies. HGFR protein is produced as a single-chain precursor, and HGF is the only known ligand. Normal HGF/HGFR signaling is essential for embryonic development, tissue repair, or wound healing, whereas aberrantly active HGFR has been strongly implicated in tumorigenesis, particularly in the development of invasive and metastatic phenotypes. HGFR protein is a multifaceted regulator of growth, motility, and invasion, and is normally expressed by cells of epithelial origin. Preclinical studies suggest that targeting aberrant HGFR signaling could be an attractive therapy in cancer.