Phosphatidylinositol 4,5-bisphosphate 3-kinases (PI3K) are a family of kinases whose activity affects pathways needed for basic cell functions. As a result, PI3K is one of the most mutated genes in all human cancers and serves as an ideal therapeutic target for cancer treatment. Expanding on work done by other groups we improved protein yield to produce stable and pure protein using a variety of modifications including improved solubility tag, novel expression modalities, and optimized purification protocol and buffer. By these means, we achieved a 40-fold increase in yield for p110α/p85α and a 3-fold increase in p110α. We also used these protocols to produce comparable constructs of the β and δ isoforms of PI3K. Increased yield enhanced the efficiency of our downstream high throughput drug discovery efforts on the PIK3 family of kinases.
Keyphrases
- drug discovery
- high throughput
- protein kinase
- single cell
- binding protein
- endothelial cells
- protein protein
- poor prognosis
- randomized controlled trial
- cell therapy
- amino acid
- genome wide
- gene expression
- small molecule
- bone marrow
- quality improvement
- transcription factor
- dna methylation
- mesenchymal stem cells
- pluripotent stem cells
- genome wide analysis