Millipore has a broad and continuously expanding portfolio of recombinant proteins. This portfolio includes more than 350 active, inactive and unactivated kinases, phosphatases and the largest available selection of disease relevant mutants. All of our enzymes are available in quantities sufficient for routine testing, as well as high throughput screening.
N-terminal GST tagged recombinant human c-Kit, amino acids 544 - end, containing the V560G mutation. The V560G mutation has been found in patients with gastrointestinal stromal tumours and has been linked to poor prognosis. This mutation confers increased susceptibility to Gleevec®.
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This gene encodes the human homolog of the proto-oncogene c-kit. C-kit was first identified as the cellular homolog of the feline sarcoma viral oncogene v-kit. This protein is a type 3 transmembrane receptor for MGF (mast cell growth factor, also known as stem cell factor). Mutations in this gene are associated with gastrointestinal stromal tumors, mast cell disease, acute myelogenous lukemia, and piebaldism. Multiple transcript variants encoding different isoforms have been found for this gene.
FUNCTION: SwissProt: P10721 # This is the receptor for stem cell factor (mast cell growth factor). It has a tyrosine-protein kinase activity. Binding of the ligands leads to the autophosphorylation of KIT and its association with substrates such as phosphatidylinositol 3-kinase (Pi3K). SIZE: 976 amino acids; 109865 Da SUBUNIT: Interacts with APS. Interacts with the tenth PDZ domain of MPDZ. SUBCELLULAR LOCATION: Membrane; Single-pass type I membrane protein. DISEASE: SwissProt: P10721 # Defects in KIT are a cause of piebaldism [MIM:172800]. Piebaldism is an autosomal dominant genetic developmental abnormality of pigmentation characterized by congenital patches of white skin and hair that lack melanocytes. & Defects in KIT are a cause of gastrointestinal stromal tumor (GIST) [MIM:606764]. & Defects in KIT have been associated with testicular tumors [MIM:273300]. It includes germ cell tumor (GCT) or testicular germ cell tumor (TGCT). SIMILARITY: SwissProt: P10721 ## Belongs to the protein kinase superfamily. Tyr protein kinase family. CSF-1/PDGF receptor subfamily. & Contains 5 Ig-like C2-type (immunoglobulin-like) domains. & Contains 1 protein kinase domain.
Stable for 6 months at -70°C from date of shipment. Upon first use, rapidly thaw the vial under cold water and immediately place on ice. Aliquot unused material into pre-chilled microcentrifuge tubes and immediately snap-freeze the vials in liquid nitrogen prior to re-storage at -70°C.
What is the best way to store and thaw enzymes?
» Answer
In order to maintain activity of enzymes, we recommend a slow thaw and aliquot storage. To do this, remove the full vial from the freezer and place under cold running tap water. Once thawed, place the vial on ice, gently mix, than make aliquots. Using liquid nitrogen or an ethanol/dry ice bath, flash freeze the aliquots. Store aliquots at the recommended temperature.
What type of buffer is best for lysis when attempting to preserve a protein?
» Answer
The best buffer is Buffer A. It is composed of 50mM Tris/HCl, pH 7.5, 1mM EGTA, 1mM EDTA, 1mM Sodium Orthovanadate (activated), 10mM Na B-glycerolphosphate, 50mM NaF, 5mM Na pyrophosphate, 1% w/v Triton X-100, 0.1% w/v 2-mercaptoethanol, 1ug/ml each of aprotinin, leupeptin and pepstatin, 1mM PMSF and 1mM microcystin (this is optional). QS with water. You may store Buffer A in aliquots at -20°C for 6 months.
Why is it important to shake the reaction mixture during the incubations?
» Answer
It is important to shake the reaction mixture during incubations so that the substrate and the enzyme get good interaction. If you do not have access to a shaking incubator, use a water bath for the incubations, removing the tubes regularly to vortex.