The availability of complete genome sequences from numerous organisms has provided investigators with the challenge of assigning physiologic functions to the encoded gene products. To facilitate this process, multiple technologies have been developed to profile the transcriptome and the proteome, including methods to monitor the function of enzymes in complex biologic systems. These methods typically target specific classes of enzymes and attempt to correlate the enzymatic activity with the specific phenotype of interest. Here, technologies to measure enzymatic activity on a subproteomic scale are reviewed, including the authors' own efforts, which are based on self-assembled microarrays utilizing peptide nucleic acid-encoded small-molecule libraries.