Modern drug development is based on the identification of target proteins that are implicated in contributing to a disease state. The protein, or proteins, are purified and used in high-throughput screening to identify potential ligands and for structural biology to elucidate structure-activity relationships in ligand-protein interactions. For this process to be successful it is essential to have a sufficient source of the target protein(s) with properties amenable to these different applications. Over the past few years, drug targets have become more complex, often constituents of multi- protein complexes or membrane associated, and this presents significant problems for obtaining sufficient purified protein with the right properties to support drug discovery
Researchers at the University of Michigan have developed a synthetic promoter, based on lac promoter, and characterized its activities. The new promoter displayed considerably lower leakage, both in overnight growth to stationary phase (14ng/ul versus 177ng/ul at 20 degrees C) and in uninduced log phase cultures. In addition, they have constructed and begun testing on a new fusion vector. Several different target genes have been cloned into this vector and put through expression trials.
Applications and Advantages
- fine-tuning of soluble expression of proteins
- broad range of responsiveness to inducer concentrations
- increasing levels of competition between the repressor and polymerase as the inducer concentration is increased