The general scheme for SHG detection in label format is shown in Figure 2. Data from a typical SHG assay is shown in Figures 3 and 4. Wild-type adenylate kinase, which catalyzes the transfer of a phosphate group from ATP to AMP, is labeled nonsite specifically with an SH-active dye and coupled to a surface.
A baseline signal is generated by the label on the protein. ATP triggers a conformational change and changes the SHG signal. A known inhibitor, AP5A, prevents a subsequent ATP-induced change. The SH signal change can either increase or decrease in a given assay, depending on whether the average orientation of the labels moves toward or away from the surface.
We have used SHG to detect ligand- and drug-induced conformational changes in a variety of protein targets such as amyloids, integrins, enzymes, and membrane receptors. We have also successfully completed several small screening projects for customers.
SHG is a break from incremental advances in drug discovery since it provides a molecular-level, functional readout in real-time. This platform is expected to be a central engine for identifying and validating both conventional and allosteric compounds, at lower cost and in cases where few other methods exist.