The inherent complexity of the proteome often demands that it be studied as manageable subsets, termed subproteomes. A subproteome can be defined in a number of ways, although a pragmatic approach is to define it based on common features in an active site that lead to binding of a common small molecule ligand (ex. a cofactor or a cross-reactive drug lead). The subproteome, so defined, can be purified using that common ligand tethered to a resin, with affinity chromatography. Affinity purification of a subproteome is described in the next chapter. That subproteome can then be analyzed using a common ligand probe, such as a fluorescent common ligand that can be used to stain members of the subproteome in a native gel. Here, we describe such a fluorescent probe, based on a catechol rhodanine acetic acid (CRAA) ligand that binds to dehydrogenases. The CRAA ligand is fluorescent and binds to dehydrogenases at pH \u3e 7, and hence can be used effectively to stain dehydrogenases in native gels to identify what subset of proteins in a mixture are dehydrogenases. Furthermore, if one is designing inhibitors to target one or more of these dehydrogenases, the CRAA staining can be performed in a competitive assay format, with or without inhibitor, to assess the selectivity of the inhibitor for the targeted dehydrogenase. Finally, the CRAA probe is a privileged scaffold for dehydrogenases, and hence can easily be modified to increase affinity for a given dehydrogenase
