Identify small molecules that inhibit kinase activity

Reference no: EM133263816

Assignment:

Vascular endothelial growth factor receptor 2 (VEGFR-2) is an endothelial cell-specific receptor that has inherent tyrosine kinase activity. The binding of the Vascular endothelial growth factor (VEGF) ligand to its receptor plays an important role in angiogenesis, a process by which new capillaries are formed. Enhanced angiogenesis is associated with a number of diseases including cancer. Your goal is to develop a homogeneous in vitro TR-FRET kinase assay for VEGFR-2 (purified protein) in an attempt to identify small molecules that inhibit kinase activity and in turn modulate angiogenesis.

Based on this background, please answer the following questions

If you wanted to measure the kinase activity of VEGFR-2, what are some of the conventional methods/assays you would use?
How would you apply a TR-FRET assay to measure the in vitro kinase activity of VEGFR-2? Describe in steps the assay design and potential readout outcome.
While setting up your experiment, you accidentally discovered autophophorylation of receptor. What observation/results led you to believe that there was autophosphorylation of the receptor?
What is the advantage of measuring autophosphorylation of the receptor rather than phosphorylation of the substrate?
How would you determine robustness of the assay and validate the assay?

Reference no: EM133263816

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